igg4 Search Results


  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 99
    Thermo Fisher igg4
    Relationship between age and immunity. Children in RTS,S vaccine group from Manhiça (black box plots) and Ilha Josina cohorts (gray box plots) were categorized into younger (12 to 24 months; Manhiça n = 11 and Ilha Josina n = 23, respectively) and older (24 to 60 months; Manhiça n = 39 and Ilha Josina n = 26, respectively) age groups. Sera collected after vaccination (month 3, M3) were tested for C1q-fixation to CSP and NANP ( a ) and <t>IgG-reactivity</t> to blood-stage antigens MSP2 and AMA1 ( b ). Samples were tested in duplicate, and the mean value was used to generate box plots for samples stratified by age group. Top, center, and bottom horizontal lines represent the 75th percentile, median, and 25th percentile, respectively; upper and lower whiskers represent the highest and lowest values within 1.5× IQR, respectively; and values that exceed this range are presented as dots. Malaria-naïve negative controls from Melbourne donors were used to calculate the seropositivity cutoff values (dashed lines), and the percentages of individuals above this threshold are shown. Reactivity between unpaired samples was compared using Mann-Whitney U test
    Igg4, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 520 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg4/product/Thermo Fisher
    Average 99 stars, based on 520 article reviews
    Price from $9.99 to $1999.99
    igg4 - by Bioz Stars, 2020-09
    99/100 stars
      Buy from Supplier

    96
    Millipore igg4
    Reverse empirical distribution function of <t>IgG3</t> levels by cross-sectional visit and treatment group in children stratified according to having (dashed line) or not (continuous line) previous episodes of clinical malaria.
    Igg4, supplied by Millipore, used in various techniques. Bioz Stars score: 96/100, based on 629 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg4/product/Millipore
    Average 96 stars, based on 629 article reviews
    Price from $9.99 to $1999.99
    igg4 - by Bioz Stars, 2020-09
    96/100 stars
      Buy from Supplier

    92
    Immunotec igg4
    Isotype responses and risk of infection: odds ratio for <t>IgG4</t> in the presence of interaction with IgG2 at P1 (solid bar) and P2 (dotted bar). The odds between the lowest IgG4 quintile (IgG4 = 1) and the highest IgG4 quintile (IgG4 = 5) are shown with 95% CI values for each IgG2 quintile. The odds were no longer significant when IgG2 levels were high (IgG2 = 4 and IgG2 = 5).
    Igg4, supplied by Immunotec, used in various techniques. Bioz Stars score: 92/100, based on 60 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg4/product/Immunotec
    Average 92 stars, based on 60 article reviews
    Price from $9.99 to $1999.99
    igg4 - by Bioz Stars, 2020-09
    92/100 stars
      Buy from Supplier

    igg4  (Abcam)
    95
    Abcam igg4
    The chronic intrathecal infusion of autoantibodies induces gait abnormalities and motor nerve conduction slowing. ( A – C ) Adult Lewis rats received daily intrathecal infusions (100 μg/d) of control <t>IgG4</t> (black circles; n = 15 animals) or anti-Nfasc155 IgG4 from patient CIDP1 (gray circles; n = 15 animals) during 3 weeks. The clinical score was monitored daily and averaged. The passive infusion of anti-Nfasc155 IgG4 induced progressive clinical symptoms. ( B and C ) Footprint analysis revealed abnormal spreading of hind limbs in animals treated with anti-Nfasc155 IgG4 compared with control animals. The footprint angle (gray lines) was significantly increased in animals treated with anti-Nfasc155 IgG4 compared with controls. ( D – G ) L6 ventral and dorsal roots from animals injected with control IgG4 or anti-Nfasc155 IgG4 were recorded on day 21 after the beginning of the injections ( n = 12–14 nerves from 12–14 animals). Representative CAPs from control IgG4–treated (black traces) and anti-Nfasc155 IgG4–treated (gray traces) animals are shown in D and F . The peak amplitude, CAP duration, and conduction velocities at peak amplitude are represented in E and G for ventral and dorsal roots, respectively. The ventral spinal nerves of animals treated with anti-Nfasc155 IgG4 showed a significant decrease in CAP amplitude and conduction velocity compared with controls. This was associated with a significant increase in CAP duration. By contrast, nerve activity was not significantly affected in dorsal root. * P
    Igg4, supplied by Abcam, used in various techniques. Bioz Stars score: 95/100, based on 79 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg4/product/Abcam
    Average 95 stars, based on 79 article reviews
    Price from $9.99 to $1999.99
    igg4 - by Bioz Stars, 2020-09
    95/100 stars
      Buy from Supplier

    93
    Becton Dickinson igg4
    Allergen-specific <t>IgG,</t> IgG 1 , IgG 4 levels during the two treatment years. (A) Phl p 1-, (B) Phl p 2-, (C) Phl p 5- and (D) Phl p 6-specific IgG (left graphs), IgG 1 and IgG 4 (right graphs) levels were measured by ELISA (Optical density levels OD 405nm , y -axes) at indicated time points ( x -axes). Lines (Placebo: red circles, BM32: blue squares) represent means with standard deviation shown. Symbols for IgG subclasses are indicated in the legends.
    Igg4, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 93/100, based on 159 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg4/product/Becton Dickinson
    Average 93 stars, based on 159 article reviews
    Price from $9.99 to $1999.99
    igg4 - by Bioz Stars, 2020-09
    93/100 stars
      Buy from Supplier

    93
    Bio-Rad igg4
    Serum <t>IgG1</t> levels of 13 children with TS before, during, and after an exacerbation of tics. Patients showed increased IgG1 levels during exacerbation: F(2, 24) = 2.63, p = .09.
    Igg4, supplied by Bio-Rad, used in various techniques. Bioz Stars score: 93/100, based on 98 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg4/product/Bio-Rad
    Average 93 stars, based on 98 article reviews
    Price from $9.99 to $1999.99
    igg4 - by Bioz Stars, 2020-09
    93/100 stars
      Buy from Supplier

    99
    Thermo Fisher igg4 levels
    Peanut specific (A) IgE, (B) <t>IgG4,</t> (C) IgE/IgG4 ratio and (D) skin prick test results at baseline and after a year of therapy for participants with proven peanut allergy (* indicates p
    Igg4 Levels, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 90 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg4 levels/product/Thermo Fisher
    Average 99 stars, based on 90 article reviews
    Price from $9.99 to $1999.99
    igg4 levels - by Bioz Stars, 2020-09
    99/100 stars
      Buy from Supplier

    94
    Novus Biologicals igg2
    <t>IgG,</t> <t>IgG1</t> and <t>IgG2a</t> antibody responses against total mixed and individual Brucella L7/L12, Omp16, Omp19 and SOD proteins in pregnant sheep (A) and goats (B) at 0, 21, 42, 63 days post-vaccination (DPV) by ELISA. Pregnant sheep and goats in the group I (Flu-BA_Omp19-SOD) were immunized twice concurrently via the subcutaneous and conjunctival routes at an interval of 21 days with vaccines generated from the influenza viral vectors (IVV) subtypes H5N1 (prime vaccination) and H1N1 (booster vaccination). The vaccination of animals of group II (Flu-BA_Omp19-SOD_TV) was carried out in the same way as in group I, but only the vaccine generated from IVV subtypes H5N1 was used, which was administered three times at 21 days intervals. Sheep and goats in the negative control group (IV) were vaccinated with 20% Montanide Gel01 adjuvant in PBS three times at 21 days intervals. Data are presented as optical density (OD) ± standard deviations; * P = 0 . 04 to P
    Igg2, supplied by Novus Biologicals, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg2/product/Novus Biologicals
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    igg2 - by Bioz Stars, 2020-09
    94/100 stars
      Buy from Supplier

    99
    Thermo Fisher igg4 antibodies
    Anti-Dsg3 autoantibodies detected in healthy blood donors do not lead to Dsg3 internalization in vitro . HaCaT cells were incubated with <t>IgG</t> from healthy individuals (NH-IgG), IgG from a pemphigus vulgaris patient (PV-IgG) or with Dsg3-positive plasma samples from healthy blood donors (samples #1382, #3167 and #9607), as well as with a Dsg3-negative sample (sample #4025). For all experimental conditions ( n = 3), the IgG concentration was 1 mg/ml (with the exception of the diluted PV-IgG samples). While NH-IgG and 4025 did not induce Dsg3 internalization, PV-IgG at 1:1 and 1:100 dilutions led to Dsg3 internalization. At low dilutions, PV-IgG and all tested Dsg3-positive samples from healthy blood donors did not show any effects on Dsg3 internalization
    Igg4 Antibodies, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 96 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg4 antibodies/product/Thermo Fisher
    Average 99 stars, based on 96 article reviews
    Price from $9.99 to $1999.99
    igg4 antibodies - by Bioz Stars, 2020-09
    99/100 stars
      Buy from Supplier

    91
    ProteomeTech Inc igg4
    Serial dilution tests performed using PROTIA™ Specific <t>IgG4®.</t> Patient sera were diluted with immunoglobulin-depleted serum. N * : negative. Specific IgG4
    Igg4, supplied by ProteomeTech Inc, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg4/product/ProteomeTech Inc
    Average 91 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    igg4 - by Bioz Stars, 2020-09
    91/100 stars
      Buy from Supplier

    92
    Cell Marque igg4
    Infectious mastoiditis mimicking <t>IgG4</t> related mastoiditis. The storiform type fibrosis is more apparent in case 4 ( a ) than case 5 ( c ). However, both cases show markedly increased numbers of IgG4 positive plasma cells ( b , d )
    Igg4, supplied by Cell Marque, used in various techniques. Bioz Stars score: 92/100, based on 33 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg4/product/Cell Marque
    Average 92 stars, based on 33 article reviews
    Price from $9.99 to $1999.99
    igg4 - by Bioz Stars, 2020-09
    92/100 stars
      Buy from Supplier

    99
    Millipore anti rabbit igg antibody
    Infectious mastoiditis mimicking <t>IgG4</t> related mastoiditis. The storiform type fibrosis is more apparent in case 4 ( a ) than case 5 ( c ). However, both cases show markedly increased numbers of IgG4 positive plasma cells ( b , d )
    Anti Rabbit Igg Antibody, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 363 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti rabbit igg antibody/product/Millipore
    Average 99 stars, based on 363 article reviews
    Price from $9.99 to $1999.99
    anti rabbit igg antibody - by Bioz Stars, 2020-09
    99/100 stars
      Buy from Supplier

    92
    Pharmingen igg4
    Whole-cell ELISA antibody levels against strain H44/76 in children with meningococcal infection, controls, siblings of children with meningococcal infection, and adults in relation to age. Means of the antibody concentration and standard errors of the means are reported for total IgG (a), IgG1 (b), IgG2 (c), IgG3 (d), <t>IgG4</t> (e), and IgM (f).
    Igg4, supplied by Pharmingen, used in various techniques. Bioz Stars score: 92/100, based on 65 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg4/product/Pharmingen
    Average 92 stars, based on 65 article reviews
    Price from $9.99 to $1999.99
    igg4 - by Bioz Stars, 2020-09
    92/100 stars
      Buy from Supplier

    94
    SouthernBiotech igg4
    HIV-specific responses and immune response capability of the case at 9.5 years of age. a Detection of HIV-specific antibodies at 9.5 years of age by western blot. The case antibody profile is compared with controls that are a high positive, low positive and HIV-negative. HIV proteins corresponding to bands in the blots are shown in the left grey-shaded block; the case profile was positive for the core proteins indicated in pink. b Quantitation of HIV-specific antibodies by multiplex bead array for all isotypes and subclasses (indicated on the left side—IgG1, IgG2, IgG3, <t>IgG4,</t> IgM, IgA1, IgA2) against gp41, Gag, RT, gp120, Nef, Tat, Vpu, peptide Vpu9 and V1V2 scaffold antigens (indicated at the top). Results are expressed as mean fluorescence intensities (MFI)), the colour key shows ranges of MFI according to colour intensity (the darker the more HIV-specific antibody detected). A result is considered positive if above the cut-off (mean ± 3 SD) determined from eight adult uninfected controls. Vpu9 amino acid sequence: STMVDMGHLRLLDVNDL. c Proportions of natural killer (NK) cells that respond to anti-CD16 antibody, and CD4+ and CD8+ T cells that respond to staphylococcal enterotoxin B (SEB) in a whole blood intracellular cytokine (ICC) assay that measures induction of interferon-γ (IFN-γ) and interleukin-2 (IL-2). HIV-uninfected adult reference values for comparison ( n = 21; median % and range)—natural killer (NK) anti-CD16%: 37.92 (12–67.6), CD4 SEB%: 6.04 (0.25–11.91), CD8 SEB%: 5.82 (0.18–18.94). d A weak positive CD4+ T cell response to Gag (0.116%) in the absence of a detectable CD8+ T cell response to Gag (
    Igg4, supplied by SouthernBiotech, used in various techniques. Bioz Stars score: 94/100, based on 279 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg4/product/SouthernBiotech
    Average 94 stars, based on 279 article reviews
    Price from $9.99 to $1999.99
    igg4 - by Bioz Stars, 2020-09
    94/100 stars
      Buy from Supplier

    94
    Meridian Life Science sd bioline onchocerciasis lf igg4 rapid test
    HIV-specific responses and immune response capability of the case at 9.5 years of age. a Detection of HIV-specific antibodies at 9.5 years of age by western blot. The case antibody profile is compared with controls that are a high positive, low positive and HIV-negative. HIV proteins corresponding to bands in the blots are shown in the left grey-shaded block; the case profile was positive for the core proteins indicated in pink. b Quantitation of HIV-specific antibodies by multiplex bead array for all isotypes and subclasses (indicated on the left side—IgG1, IgG2, IgG3, <t>IgG4,</t> IgM, IgA1, IgA2) against gp41, Gag, RT, gp120, Nef, Tat, Vpu, peptide Vpu9 and V1V2 scaffold antigens (indicated at the top). Results are expressed as mean fluorescence intensities (MFI)), the colour key shows ranges of MFI according to colour intensity (the darker the more HIV-specific antibody detected). A result is considered positive if above the cut-off (mean ± 3 SD) determined from eight adult uninfected controls. Vpu9 amino acid sequence: STMVDMGHLRLLDVNDL. c Proportions of natural killer (NK) cells that respond to anti-CD16 antibody, and CD4+ and CD8+ T cells that respond to staphylococcal enterotoxin B (SEB) in a whole blood intracellular cytokine (ICC) assay that measures induction of interferon-γ (IFN-γ) and interleukin-2 (IL-2). HIV-uninfected adult reference values for comparison ( n = 21; median % and range)—natural killer (NK) anti-CD16%: 37.92 (12–67.6), CD4 SEB%: 6.04 (0.25–11.91), CD8 SEB%: 5.82 (0.18–18.94). d A weak positive CD4+ T cell response to Gag (0.116%) in the absence of a detectable CD8+ T cell response to Gag (
    Sd Bioline Onchocerciasis Lf Igg4 Rapid Test, supplied by Meridian Life Science, used in various techniques. Bioz Stars score: 94/100, based on 24 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/sd bioline onchocerciasis lf igg4 rapid test/product/Meridian Life Science
    Average 94 stars, based on 24 article reviews
    Price from $9.99 to $1999.99
    sd bioline onchocerciasis lf igg4 rapid test - by Bioz Stars, 2020-09
    94/100 stars
      Buy from Supplier

    96
    Thermo Fisher anti rabbit igg
    A strong correlation between autoantibody binding to <t>IgG</t> complexed with each <t>HLA-DR</t> allele and the odds ratio for that allele’s association with RA. ( A ) The IgGH was cotransfected with Ig light chain (L), Ii, and HLA-DRα in combination
    Anti Rabbit Igg, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 96/100, based on 7812 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti rabbit igg/product/Thermo Fisher
    Average 96 stars, based on 7812 article reviews
    Price from $9.99 to $1999.99
    anti rabbit igg - by Bioz Stars, 2020-09
    96/100 stars
      Buy from Supplier

    95
    Millipore anti human igg4 biotin antibody mouse monoclonal
    Correlation of the effusion κ/λ free L chains (FLC) ratio with immunoglobulin (Ig)G4 + plasma cell counts (a) and <t>IgG4</t> + /IgG + plasma cell ratio (b) in the pleura of patients in the IgG4 + group. *One‐tailed P ‐value.
    Anti Human Igg4 Biotin Antibody Mouse Monoclonal, supplied by Millipore, used in various techniques. Bioz Stars score: 95/100, based on 54 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti human igg4 biotin antibody mouse monoclonal/product/Millipore
    Average 95 stars, based on 54 article reviews
    Price from $9.99 to $1999.99
    anti human igg4 biotin antibody mouse monoclonal - by Bioz Stars, 2020-09
    95/100 stars
      Buy from Supplier

    99
    Millipore human igg4
    (A) Fab fragments from FS-1 <t>IgG4</t> bind wDsg1(blue), bind partially to Dsg1-m1 (red) and do not bind Dsg1-m2 (green) or to wDsg4 (purple) that overlap in a single line. (B) Same Fab fragments from FS1 IgG4 are pathogenic when tested on the mouse model. The animals show skin blisters, which are subcorneal and the Fab fragments are bound to the surface of keratinocytes undergoing detachment at the site of injury by direct immunofluorescence (IF). Orignal magnification of histology and direct IF, x200.
    Human Igg4, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 92 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/human igg4/product/Millipore
    Average 99 stars, based on 92 article reviews
    Price from $9.99 to $1999.99
    human igg4 - by Bioz Stars, 2020-09
    99/100 stars
      Buy from Supplier

    99
    Thermo Fisher mouse anti human igg4 fc secondary antibody
    Onchocerca volvulus adult worm (OvAg) and recombinant Ov16 antigen-specific <t>IgG4</t> reactivity (optical densities; OD) in participants and positive and negative IgG4 responses in age groups. In A and C the data on antigen-specific IgG4 reactivity are shown as mean optical densities (ODs) with 95% confidence intervals for the means (diamonds). The data presented in box plots show the median OD per age group with the 25% and 75% quartiles and the 1.5x of the interquartile range. In B and D the antigen-specific-IgG4 positive and negative responses in age groups are indicated (in %).
    Mouse Anti Human Igg4 Fc Secondary Antibody, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 18 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse anti human igg4 fc secondary antibody/product/Thermo Fisher
    Average 99 stars, based on 18 article reviews
    Price from $9.99 to $1999.99
    mouse anti human igg4 fc secondary antibody - by Bioz Stars, 2020-09
    99/100 stars
      Buy from Supplier

    93
    Millipore anti human igg4 biotin antibody
    Evaluating <t>IgG</t> subclass response to the 39041 epitope ( n = 6). The Kruskal-Wallis test was used for analyzing differences between each IgG subclass response in P. vivax -exposed individuals' samples. ** p
    Anti Human Igg4 Biotin Antibody, supplied by Millipore, used in various techniques. Bioz Stars score: 93/100, based on 16 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti human igg4 biotin antibody/product/Millipore
    Average 93 stars, based on 16 article reviews
    Price from $9.99 to $1999.99
    anti human igg4 biotin antibody - by Bioz Stars, 2020-09
    93/100 stars
      Buy from Supplier

    92
    Millipore anti igg
    Evaluating <t>IgG</t> subclass response to the 39041 epitope ( n = 6). The Kruskal-Wallis test was used for analyzing differences between each IgG subclass response in P. vivax -exposed individuals' samples. ** p
    Anti Igg, supplied by Millipore, used in various techniques. Bioz Stars score: 92/100, based on 529 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti igg/product/Millipore
    Average 92 stars, based on 529 article reviews
    Price from $9.99 to $1999.99
    anti igg - by Bioz Stars, 2020-09
    92/100 stars
      Buy from Supplier

    Image Search Results


    Relationship between age and immunity. Children in RTS,S vaccine group from Manhiça (black box plots) and Ilha Josina cohorts (gray box plots) were categorized into younger (12 to 24 months; Manhiça n = 11 and Ilha Josina n = 23, respectively) and older (24 to 60 months; Manhiça n = 39 and Ilha Josina n = 26, respectively) age groups. Sera collected after vaccination (month 3, M3) were tested for C1q-fixation to CSP and NANP ( a ) and IgG-reactivity to blood-stage antigens MSP2 and AMA1 ( b ). Samples were tested in duplicate, and the mean value was used to generate box plots for samples stratified by age group. Top, center, and bottom horizontal lines represent the 75th percentile, median, and 25th percentile, respectively; upper and lower whiskers represent the highest and lowest values within 1.5× IQR, respectively; and values that exceed this range are presented as dots. Malaria-naïve negative controls from Melbourne donors were used to calculate the seropositivity cutoff values (dashed lines), and the percentages of individuals above this threshold are shown. Reactivity between unpaired samples was compared using Mann-Whitney U test

    Journal: BMC Medicine

    Article Title: Induction and decay of functional complement-fixing antibodies by the RTS,S malaria vaccine in children, and a negative impact of malaria exposure

    doi: 10.1186/s12916-019-1277-x

    Figure Lengend Snippet: Relationship between age and immunity. Children in RTS,S vaccine group from Manhiça (black box plots) and Ilha Josina cohorts (gray box plots) were categorized into younger (12 to 24 months; Manhiça n = 11 and Ilha Josina n = 23, respectively) and older (24 to 60 months; Manhiça n = 39 and Ilha Josina n = 26, respectively) age groups. Sera collected after vaccination (month 3, M3) were tested for C1q-fixation to CSP and NANP ( a ) and IgG-reactivity to blood-stage antigens MSP2 and AMA1 ( b ). Samples were tested in duplicate, and the mean value was used to generate box plots for samples stratified by age group. Top, center, and bottom horizontal lines represent the 75th percentile, median, and 25th percentile, respectively; upper and lower whiskers represent the highest and lowest values within 1.5× IQR, respectively; and values that exceed this range are presented as dots. Malaria-naïve negative controls from Melbourne donors were used to calculate the seropositivity cutoff values (dashed lines), and the percentages of individuals above this threshold are shown. Reactivity between unpaired samples was compared using Mann-Whitney U test

    Article Snippet: To detect IgG subclasses, plates were instead incubated with mouse anti-human IgG1, IgG2, IgG3, and IgG4 (Thermo Fisher Scientific), washed, and then incubated with goat anti-mouse IgG HRP (Millipore) all at 1/1000 dilution in buffer for 1 h at RT.

    Techniques: MANN-WHITNEY

    RTS,S vaccine-induced immunity declines over time. A random selection of children vaccinated with RTS,S (Manhiça cohort, n = 30) was tested for C1q-fixation ( a ), IgG/IgM ( b ), and IgG subclasses ( c ) to CSP at months 3, 8.5, 21, 33, 45, and 6. Note that due to low reactivity, C1q-fixation was re-tested at a higher dilution of 1/110, in addition to 1/250, to confirm results. Samples were tested in duplicate, and the median and 95% CI of the median from each time point group are shown by the symbol and shaded area, respectively

    Journal: BMC Medicine

    Article Title: Induction and decay of functional complement-fixing antibodies by the RTS,S malaria vaccine in children, and a negative impact of malaria exposure

    doi: 10.1186/s12916-019-1277-x

    Figure Lengend Snippet: RTS,S vaccine-induced immunity declines over time. A random selection of children vaccinated with RTS,S (Manhiça cohort, n = 30) was tested for C1q-fixation ( a ), IgG/IgM ( b ), and IgG subclasses ( c ) to CSP at months 3, 8.5, 21, 33, 45, and 6. Note that due to low reactivity, C1q-fixation was re-tested at a higher dilution of 1/110, in addition to 1/250, to confirm results. Samples were tested in duplicate, and the median and 95% CI of the median from each time point group are shown by the symbol and shaded area, respectively

    Article Snippet: To detect IgG subclasses, plates were instead incubated with mouse anti-human IgG1, IgG2, IgG3, and IgG4 (Thermo Fisher Scientific), washed, and then incubated with goat anti-mouse IgG HRP (Millipore) all at 1/1000 dilution in buffer for 1 h at RT.

    Techniques: Selection

    RTS,S vaccine-induced IgG and IgM antibodies to CSP. Children in RTS,S and comparator vaccine groups from Manhiça (black box plots; N = 50 and N = 25, respectively) and Ilha Josina cohorts (gray box plots; N = 49 and N = 24, respectively) were tested for IgG ( a ), IgG subclasses ( b ), and IgM ( c ) to CSP. Sera collected at baseline (month 0, M0) and after vaccination (month 3, M3) were tested in duplicate (note that only M3 was tested in b ), and the mean value was used to generate box plots whereby top, center, and bottom horizontal lines represent the 75th percentile, median, and 25th percentile, respectively; upper and lower whiskers represent the highest and lowest values within 1.5× IQR, respectively; and values that exceed this range are presented as dots. Malaria-naïve negative controls from Melbourne donors were used to calculate the seropositivity cutoff value (dashed lines), and the percentage of individuals above this threshold are shown. Reactivity between paired samples and unpaired samples were compared using Wilcoxon matched-pairs signed-rank test and Mann-Whitney U test, respectively

    Journal: BMC Medicine

    Article Title: Induction and decay of functional complement-fixing antibodies by the RTS,S malaria vaccine in children, and a negative impact of malaria exposure

    doi: 10.1186/s12916-019-1277-x

    Figure Lengend Snippet: RTS,S vaccine-induced IgG and IgM antibodies to CSP. Children in RTS,S and comparator vaccine groups from Manhiça (black box plots; N = 50 and N = 25, respectively) and Ilha Josina cohorts (gray box plots; N = 49 and N = 24, respectively) were tested for IgG ( a ), IgG subclasses ( b ), and IgM ( c ) to CSP. Sera collected at baseline (month 0, M0) and after vaccination (month 3, M3) were tested in duplicate (note that only M3 was tested in b ), and the mean value was used to generate box plots whereby top, center, and bottom horizontal lines represent the 75th percentile, median, and 25th percentile, respectively; upper and lower whiskers represent the highest and lowest values within 1.5× IQR, respectively; and values that exceed this range are presented as dots. Malaria-naïve negative controls from Melbourne donors were used to calculate the seropositivity cutoff value (dashed lines), and the percentage of individuals above this threshold are shown. Reactivity between paired samples and unpaired samples were compared using Wilcoxon matched-pairs signed-rank test and Mann-Whitney U test, respectively

    Article Snippet: To detect IgG subclasses, plates were instead incubated with mouse anti-human IgG1, IgG2, IgG3, and IgG4 (Thermo Fisher Scientific), washed, and then incubated with goat anti-mouse IgG HRP (Millipore) all at 1/1000 dilution in buffer for 1 h at RT.

    Techniques: MANN-WHITNEY

    High variability among RTS,S vaccine-induced IgG targeting the central repeat and C-terminal regions of CSP. Children in RTS,S vaccine group from Manhiça and Ilha Josina cohorts ( N = 99, 3 M) were tested for IgG to NANP and CT and C1q-fixation to CSP, and the values were used for the following analysis. a Heat map of children arranged in descending (top to bottom) order of C1q-fixation (left), corresponding IgG to NANP and CT (middle), and for comparison IgM to NANP and CT (right). b Variability between epitope specificity was quantified by calculating the ratio of NANP-to-CT IgG. Children with low variability were considered to have equal reactivity to NANP and CT (ratio between 0.75 and 1.25 shown in white), and children exceeding this range were considered to have a NANP- or CT-skewed response (ratio > 1.25 shown in purple and ratio

    Journal: BMC Medicine

    Article Title: Induction and decay of functional complement-fixing antibodies by the RTS,S malaria vaccine in children, and a negative impact of malaria exposure

    doi: 10.1186/s12916-019-1277-x

    Figure Lengend Snippet: High variability among RTS,S vaccine-induced IgG targeting the central repeat and C-terminal regions of CSP. Children in RTS,S vaccine group from Manhiça and Ilha Josina cohorts ( N = 99, 3 M) were tested for IgG to NANP and CT and C1q-fixation to CSP, and the values were used for the following analysis. a Heat map of children arranged in descending (top to bottom) order of C1q-fixation (left), corresponding IgG to NANP and CT (middle), and for comparison IgM to NANP and CT (right). b Variability between epitope specificity was quantified by calculating the ratio of NANP-to-CT IgG. Children with low variability were considered to have equal reactivity to NANP and CT (ratio between 0.75 and 1.25 shown in white), and children exceeding this range were considered to have a NANP- or CT-skewed response (ratio > 1.25 shown in purple and ratio

    Article Snippet: To detect IgG subclasses, plates were instead incubated with mouse anti-human IgG1, IgG2, IgG3, and IgG4 (Thermo Fisher Scientific), washed, and then incubated with goat anti-mouse IgG HRP (Millipore) all at 1/1000 dilution in buffer for 1 h at RT.

    Techniques:

    Functional complement-fixing antibodies target the central repeat and C-terminal regions of CSP. Children in RTS,S vaccine group from Manhiça (black box plots; N = 50) and Ilha Josina cohorts (gray box plots; N = 49) were tested for IgG ( a ) and IgM ( b ) to NANP and CT regions of CSP. Sera collected at baseline (month 0, M0) and after vaccination (month 3, M3) were tested in duplicate, and the mean value was used to generate box plots whereby top, center, and bottom horizontal lines represent the 75th percentile, median, and 25th percentile, respectively; upper and lower whiskers represent the highest and lowest values within 1.5× IQR, respectively; and values that exceed this range are presented as dots. Malaria-naïve negative controls from Melbourne donors were used to calculate the seropositivity cutoff values (dashed lines), and the percentages of individuals above this threshold are shown. Reactivity between paired samples was compared using Wilcoxon matched-pairs signed-rank test. c , d Children in RTS,S vaccine group from Manhiça and Ilha Josina cohorts ( N = 99, M3) were tested for C1q-fixation to CSP, NANP, and CT, and the values were plotted compared to IgG reactivity (c) and C1q fixation to CSP, NANP, and CT were correlated (d)

    Journal: BMC Medicine

    Article Title: Induction and decay of functional complement-fixing antibodies by the RTS,S malaria vaccine in children, and a negative impact of malaria exposure

    doi: 10.1186/s12916-019-1277-x

    Figure Lengend Snippet: Functional complement-fixing antibodies target the central repeat and C-terminal regions of CSP. Children in RTS,S vaccine group from Manhiça (black box plots; N = 50) and Ilha Josina cohorts (gray box plots; N = 49) were tested for IgG ( a ) and IgM ( b ) to NANP and CT regions of CSP. Sera collected at baseline (month 0, M0) and after vaccination (month 3, M3) were tested in duplicate, and the mean value was used to generate box plots whereby top, center, and bottom horizontal lines represent the 75th percentile, median, and 25th percentile, respectively; upper and lower whiskers represent the highest and lowest values within 1.5× IQR, respectively; and values that exceed this range are presented as dots. Malaria-naïve negative controls from Melbourne donors were used to calculate the seropositivity cutoff values (dashed lines), and the percentages of individuals above this threshold are shown. Reactivity between paired samples was compared using Wilcoxon matched-pairs signed-rank test. c , d Children in RTS,S vaccine group from Manhiça and Ilha Josina cohorts ( N = 99, M3) were tested for C1q-fixation to CSP, NANP, and CT, and the values were plotted compared to IgG reactivity (c) and C1q fixation to CSP, NANP, and CT were correlated (d)

    Article Snippet: To detect IgG subclasses, plates were instead incubated with mouse anti-human IgG1, IgG2, IgG3, and IgG4 (Thermo Fisher Scientific), washed, and then incubated with goat anti-mouse IgG HRP (Millipore) all at 1/1000 dilution in buffer for 1 h at RT.

    Techniques: Functional Assay

    Reverse empirical distribution function of IgG3 levels by cross-sectional visit and treatment group in children stratified according to having (dashed line) or not (continuous line) previous episodes of clinical malaria.

    Journal: Malaria Journal

    Article Title: Impact of age of first exposure to Plasmodium falciparum on antibody responses to malaria in children: a randomized, controlled trial in Mozambique

    doi: 10.1186/1475-2875-13-121

    Figure Lengend Snippet: Reverse empirical distribution function of IgG3 levels by cross-sectional visit and treatment group in children stratified according to having (dashed line) or not (continuous line) previous episodes of clinical malaria.

    Article Snippet: After one-hour incubation with 100 μL of peroxidase-conjugated rabbit anti-human IgG (1/8,000), IgM (1/3,000) (DakoCytomation) and sheep anti-human IgG1 (1/6,000), IgG2 (1/1,500), IgG3 (1/1,500), IgG4 (1/3,000) isotypes (Binding site, UK) secondary antibodies diluted in wash buffer, respectively, 100 μL of citric acid 0.243 M + Na2 HPO4 0.512 M + 1 mg/ml of o-phenylenediamine chromogen (Sigma, St Louis, MO, USA) with 0.012% H2 O2 substrate were added per well for 5 min, and the colorimetric reaction was stopped with 25 μL of 3 M H2 SO4 .

    Techniques:

    Weighted scatter plots of IgG levels by cross-sectional visit and treatment group. Red symbols correspond to IgG levels in those children with previous or current Plasmodium falciparum infection.

    Journal: Malaria Journal

    Article Title: Impact of age of first exposure to Plasmodium falciparum on antibody responses to malaria in children: a randomized, controlled trial in Mozambique

    doi: 10.1186/1475-2875-13-121

    Figure Lengend Snippet: Weighted scatter plots of IgG levels by cross-sectional visit and treatment group. Red symbols correspond to IgG levels in those children with previous or current Plasmodium falciparum infection.

    Article Snippet: After one-hour incubation with 100 μL of peroxidase-conjugated rabbit anti-human IgG (1/8,000), IgM (1/3,000) (DakoCytomation) and sheep anti-human IgG1 (1/6,000), IgG2 (1/1,500), IgG3 (1/1,500), IgG4 (1/3,000) isotypes (Binding site, UK) secondary antibodies diluted in wash buffer, respectively, 100 μL of citric acid 0.243 M + Na2 HPO4 0.512 M + 1 mg/ml of o-phenylenediamine chromogen (Sigma, St Louis, MO, USA) with 0.012% H2 O2 substrate were added per well for 5 min, and the colorimetric reaction was stopped with 25 μL of 3 M H2 SO4 .

    Techniques: Infection

    Reverse empirical distribution function of IgG levels by cross-sectional visit and treatment group in children stratified according to having (dashed line) or not (continuous line) previous episodes of clinical malaria.

    Journal: Malaria Journal

    Article Title: Impact of age of first exposure to Plasmodium falciparum on antibody responses to malaria in children: a randomized, controlled trial in Mozambique

    doi: 10.1186/1475-2875-13-121

    Figure Lengend Snippet: Reverse empirical distribution function of IgG levels by cross-sectional visit and treatment group in children stratified according to having (dashed line) or not (continuous line) previous episodes of clinical malaria.

    Article Snippet: After one-hour incubation with 100 μL of peroxidase-conjugated rabbit anti-human IgG (1/8,000), IgM (1/3,000) (DakoCytomation) and sheep anti-human IgG1 (1/6,000), IgG2 (1/1,500), IgG3 (1/1,500), IgG4 (1/3,000) isotypes (Binding site, UK) secondary antibodies diluted in wash buffer, respectively, 100 μL of citric acid 0.243 M + Na2 HPO4 0.512 M + 1 mg/ml of o-phenylenediamine chromogen (Sigma, St Louis, MO, USA) with 0.012% H2 O2 substrate were added per well for 5 min, and the colorimetric reaction was stopped with 25 μL of 3 M H2 SO4 .

    Techniques:

    Reverse empirical distribution function of IgG1 levels by cross-sectional visit and treatment group in children stratified according to having (dashed line) or not (continuous line) previous episodes of clinical malaria.

    Journal: Malaria Journal

    Article Title: Impact of age of first exposure to Plasmodium falciparum on antibody responses to malaria in children: a randomized, controlled trial in Mozambique

    doi: 10.1186/1475-2875-13-121

    Figure Lengend Snippet: Reverse empirical distribution function of IgG1 levels by cross-sectional visit and treatment group in children stratified according to having (dashed line) or not (continuous line) previous episodes of clinical malaria.

    Article Snippet: After one-hour incubation with 100 μL of peroxidase-conjugated rabbit anti-human IgG (1/8,000), IgM (1/3,000) (DakoCytomation) and sheep anti-human IgG1 (1/6,000), IgG2 (1/1,500), IgG3 (1/1,500), IgG4 (1/3,000) isotypes (Binding site, UK) secondary antibodies diluted in wash buffer, respectively, 100 μL of citric acid 0.243 M + Na2 HPO4 0.512 M + 1 mg/ml of o-phenylenediamine chromogen (Sigma, St Louis, MO, USA) with 0.012% H2 O2 substrate were added per well for 5 min, and the colorimetric reaction was stopped with 25 μL of 3 M H2 SO4 .

    Techniques:

    Detection of IgM and IgG antibodies against LPS O26 in the serum from patients with HUS and controls by ELISA. Serum samples were diluted 1:500 in PBS-Tween, and absorbance values (Abs) were measured at 492 nm.

    Journal: The Open Microbiology Journal

    Article Title: Hemolytic Uremic Syndrome in Pediatric Intensive Care Units in S?o Paulo, Brazil

    doi: 10.2174/1874285801105010076

    Figure Lengend Snippet: Detection of IgM and IgG antibodies against LPS O26 in the serum from patients with HUS and controls by ELISA. Serum samples were diluted 1:500 in PBS-Tween, and absorbance values (Abs) were measured at 492 nm.

    Article Snippet: Presence of IgM and IgG antibodies was investigated in the samples by using anti-human IgM and IgG conjugated peroxidase (Sigma) diluted 1:1000 and incubated for 2 hours at room temperature.

    Techniques: Enzyme-linked Immunosorbent Assay

    Detection of. IgM and IgG antibodies against LPS O111 in the serum from patients with HUS and controls by ELISA. Serum samples were diluted 1:500 in PBS-Tween, and absorbance values (Abs) were measured at 492 nm.

    Journal: The Open Microbiology Journal

    Article Title: Hemolytic Uremic Syndrome in Pediatric Intensive Care Units in S?o Paulo, Brazil

    doi: 10.2174/1874285801105010076

    Figure Lengend Snippet: Detection of. IgM and IgG antibodies against LPS O111 in the serum from patients with HUS and controls by ELISA. Serum samples were diluted 1:500 in PBS-Tween, and absorbance values (Abs) were measured at 492 nm.

    Article Snippet: Presence of IgM and IgG antibodies was investigated in the samples by using anti-human IgM and IgG conjugated peroxidase (Sigma) diluted 1:1000 and incubated for 2 hours at room temperature.

    Techniques: Enzyme-linked Immunosorbent Assay

    Detection of IgM and IgG antibodies against LPS O157 in the serum from patients with HUS and controls by ELISA. Serum samples were diluted 1:500 in PBS-Tween, and absorbance values (Abs) were measured at 492 nm.

    Journal: The Open Microbiology Journal

    Article Title: Hemolytic Uremic Syndrome in Pediatric Intensive Care Units in S?o Paulo, Brazil

    doi: 10.2174/1874285801105010076

    Figure Lengend Snippet: Detection of IgM and IgG antibodies against LPS O157 in the serum from patients with HUS and controls by ELISA. Serum samples were diluted 1:500 in PBS-Tween, and absorbance values (Abs) were measured at 492 nm.

    Article Snippet: Presence of IgM and IgG antibodies was investigated in the samples by using anti-human IgM and IgG conjugated peroxidase (Sigma) diluted 1:1000 and incubated for 2 hours at room temperature.

    Techniques: Enzyme-linked Immunosorbent Assay

    Isotype responses and risk of infection: odds ratio for IgG4 in the presence of interaction with IgG2 at P1 (solid bar) and P2 (dotted bar). The odds between the lowest IgG4 quintile (IgG4 = 1) and the highest IgG4 quintile (IgG4 = 5) are shown with 95% CI values for each IgG2 quintile. The odds were no longer significant when IgG2 levels were high (IgG2 = 4 and IgG2 = 5).

    Journal: Infection and Immunity

    Article Title: High Immunoglobulin G2 (IgG2) and Low IgG4 Levels Are Associated with Human Resistance to Plasmodium falciparum Malaria

    doi:

    Figure Lengend Snippet: Isotype responses and risk of infection: odds ratio for IgG4 in the presence of interaction with IgG2 at P1 (solid bar) and P2 (dotted bar). The odds between the lowest IgG4 quintile (IgG4 = 1) and the highest IgG4 quintile (IgG4 = 5) are shown with 95% CI values for each IgG2 quintile. The odds were no longer significant when IgG2 levels were high (IgG2 = 4 and IgG2 = 5).

    Article Snippet: Since interaction between IgG2 and IgG4 was significant at P2 ( P < 0.0001), the odds ratio depended on IgG2 levels (Table and Fig. ).

    Techniques: Infection

    The chronic intrathecal infusion of autoantibodies induces gait abnormalities and motor nerve conduction slowing. ( A – C ) Adult Lewis rats received daily intrathecal infusions (100 μg/d) of control IgG4 (black circles; n = 15 animals) or anti-Nfasc155 IgG4 from patient CIDP1 (gray circles; n = 15 animals) during 3 weeks. The clinical score was monitored daily and averaged. The passive infusion of anti-Nfasc155 IgG4 induced progressive clinical symptoms. ( B and C ) Footprint analysis revealed abnormal spreading of hind limbs in animals treated with anti-Nfasc155 IgG4 compared with control animals. The footprint angle (gray lines) was significantly increased in animals treated with anti-Nfasc155 IgG4 compared with controls. ( D – G ) L6 ventral and dorsal roots from animals injected with control IgG4 or anti-Nfasc155 IgG4 were recorded on day 21 after the beginning of the injections ( n = 12–14 nerves from 12–14 animals). Representative CAPs from control IgG4–treated (black traces) and anti-Nfasc155 IgG4–treated (gray traces) animals are shown in D and F . The peak amplitude, CAP duration, and conduction velocities at peak amplitude are represented in E and G for ventral and dorsal roots, respectively. The ventral spinal nerves of animals treated with anti-Nfasc155 IgG4 showed a significant decrease in CAP amplitude and conduction velocity compared with controls. This was associated with a significant increase in CAP duration. By contrast, nerve activity was not significantly affected in dorsal root. * P

    Journal: The Journal of Clinical Investigation

    Article Title: Anti–neurofascin-155 IgG4 antibodies prevent paranodal complex formation in vivo

    doi: 10.1172/JCI124694

    Figure Lengend Snippet: The chronic intrathecal infusion of autoantibodies induces gait abnormalities and motor nerve conduction slowing. ( A – C ) Adult Lewis rats received daily intrathecal infusions (100 μg/d) of control IgG4 (black circles; n = 15 animals) or anti-Nfasc155 IgG4 from patient CIDP1 (gray circles; n = 15 animals) during 3 weeks. The clinical score was monitored daily and averaged. The passive infusion of anti-Nfasc155 IgG4 induced progressive clinical symptoms. ( B and C ) Footprint analysis revealed abnormal spreading of hind limbs in animals treated with anti-Nfasc155 IgG4 compared with control animals. The footprint angle (gray lines) was significantly increased in animals treated with anti-Nfasc155 IgG4 compared with controls. ( D – G ) L6 ventral and dorsal roots from animals injected with control IgG4 or anti-Nfasc155 IgG4 were recorded on day 21 after the beginning of the injections ( n = 12–14 nerves from 12–14 animals). Representative CAPs from control IgG4–treated (black traces) and anti-Nfasc155 IgG4–treated (gray traces) animals are shown in D and F . The peak amplitude, CAP duration, and conduction velocities at peak amplitude are represented in E and G for ventral and dorsal roots, respectively. The ventral spinal nerves of animals treated with anti-Nfasc155 IgG4 showed a significant decrease in CAP amplitude and conduction velocity compared with controls. This was associated with a significant increase in CAP duration. By contrast, nerve activity was not significantly affected in dorsal root. * P

    Article Snippet: Purified antibody samples were denatured in SDS sample buffer for 2 minutes at 92°C, loaded on 7.5% SDS-PAGE gels, transferred, and immunoblotted with mouse monoclonal antibodies against human IgG1 (1:1000; 4E3, Abcam), IgG2 (1:1000; HP6014, Abcam), IgG3 (1:1000; HP-6050, Sigma-Aldrich), or IgG4 (1:5000; HP6035, Abcam).

    Techniques: Injection, Activity Assay

    Autoantibodies induce the selective depletion of Nfasc155 in ventral spinal nerve. ( A ) Ventral and dorsal spinal nerve proteins (250 μg) from adult animals injected with control IgG4 ( n = 4–6) or anti-Nfasc155 IgG4 from patient CIDP1 ( n = 4–6) for 3 weeks were separated on SDS-PAGE gels and immunoblotted with antibodies against Nfasc155, CASPR1, neurofilament-200, and E-cadherin. Arrowheads indicate Nfasc155-H and Nfasc155-L isoforms. ( B ) The levels of Nfasc155, CASPR1, neurofilament-200, and E-cadherin were evaluated in animals treated with anti-Nfasc155 IgG4 relatively to those in control animals in both ventral and dorsal spinal roots. Nfasc155 levels were significantly decreased in ventral spinal nerves, but not in dorsal spinal nerves (* P

    Journal: The Journal of Clinical Investigation

    Article Title: Anti–neurofascin-155 IgG4 antibodies prevent paranodal complex formation in vivo

    doi: 10.1172/JCI124694

    Figure Lengend Snippet: Autoantibodies induce the selective depletion of Nfasc155 in ventral spinal nerve. ( A ) Ventral and dorsal spinal nerve proteins (250 μg) from adult animals injected with control IgG4 ( n = 4–6) or anti-Nfasc155 IgG4 from patient CIDP1 ( n = 4–6) for 3 weeks were separated on SDS-PAGE gels and immunoblotted with antibodies against Nfasc155, CASPR1, neurofilament-200, and E-cadherin. Arrowheads indicate Nfasc155-H and Nfasc155-L isoforms. ( B ) The levels of Nfasc155, CASPR1, neurofilament-200, and E-cadherin were evaluated in animals treated with anti-Nfasc155 IgG4 relatively to those in control animals in both ventral and dorsal spinal roots. Nfasc155 levels were significantly decreased in ventral spinal nerves, but not in dorsal spinal nerves (* P

    Article Snippet: Purified antibody samples were denatured in SDS sample buffer for 2 minutes at 92°C, loaded on 7.5% SDS-PAGE gels, transferred, and immunoblotted with mouse monoclonal antibodies against human IgG1 (1:1000; 4E3, Abcam), IgG2 (1:1000; HP6014, Abcam), IgG3 (1:1000; HP-6050, Sigma-Aldrich), or IgG4 (1:5000; HP6035, Abcam).

    Techniques: Injection, SDS Page

    Autoantibodies preferentially affect motor axons. ( A – D ) The distribution of the axonal population according to the nodal diameters of axons was examined in L6 ventral roots ( A and B ) and dorsal roots ( C and D ) of animals treated with control IgG4 ( n = 1297 nodes for ventral roots and 1061 nodes for dorsal roots from 11 animals) or anti-Nfasc155 IgG4 from patient CIDP1 ( n = 1376 nodes for ventral roots and 875 nodes for dorsal roots from 11 animals). The respective proportion of normal nodes (open boxes) or nodes lacking paranodes (hatched boxes) is represented on the left, and the distribution of the total axonal population is represented on the right. The percentage of nodes showing paranodal alterations was calculated in ventral ( B ) and dorsal roots ( D ). Scatter plots represent the percentage of normal nodes in each animal. Paranodes were more significantly affected in ventral root axons than in dorsal roots (** P

    Journal: The Journal of Clinical Investigation

    Article Title: Anti–neurofascin-155 IgG4 antibodies prevent paranodal complex formation in vivo

    doi: 10.1172/JCI124694

    Figure Lengend Snippet: Autoantibodies preferentially affect motor axons. ( A – D ) The distribution of the axonal population according to the nodal diameters of axons was examined in L6 ventral roots ( A and B ) and dorsal roots ( C and D ) of animals treated with control IgG4 ( n = 1297 nodes for ventral roots and 1061 nodes for dorsal roots from 11 animals) or anti-Nfasc155 IgG4 from patient CIDP1 ( n = 1376 nodes for ventral roots and 875 nodes for dorsal roots from 11 animals). The respective proportion of normal nodes (open boxes) or nodes lacking paranodes (hatched boxes) is represented on the left, and the distribution of the total axonal population is represented on the right. The percentage of nodes showing paranodal alterations was calculated in ventral ( B ) and dorsal roots ( D ). Scatter plots represent the percentage of normal nodes in each animal. Paranodes were more significantly affected in ventral root axons than in dorsal roots (** P

    Article Snippet: Purified antibody samples were denatured in SDS sample buffer for 2 minutes at 92°C, loaded on 7.5% SDS-PAGE gels, transferred, and immunoblotted with mouse monoclonal antibodies against human IgG1 (1:1000; 4E3, Abcam), IgG2 (1:1000; HP6014, Abcam), IgG3 (1:1000; HP-6050, Sigma-Aldrich), or IgG4 (1:5000; HP6035, Abcam).

    Techniques:

    Schematic representation of the pathogenic mechanisms of anti–paranodal protein autoantibodies. ( A ) Representation of a mature node of Ranvier and myelinated axon. Myelin (yellow) covers the axon except at the node of Ranvier (red). At paranodal junctions (green), Nfasc155 interacts with its axonal partners CASPR1/CNTN1. Nfasc155 is also found at Schmidt-Lanterman incisures (dashed green lines). The Schwann cell nucleus is shown in gray. ( B ) In neonatal animals, the progressive enwrapping of axons by Schwann cells induces the formation of paranodal regions at node borders. The injection of anti-Nfasc155 IgG4 (blue) during the neonatal period does not affect myelination or node/paranode formation, but induces the depletion of Nfasc155, and thereby alters the formation of paranodal septate-like junctions. ( C ) At adult age, evidence suggests that the Nfasc155/CASPR1/CNTN1 complex is constantly renewed at paranodes possibly through degradation and replenishment mechanisms. The chronic infusion of anti-Nfasc155 IgG4 (blue) may preclude the regeneration of the paranodal axoglial junction by inducing Nfasc155 depletion, and thereby alter paranode structure and conduction.

    Journal: The Journal of Clinical Investigation

    Article Title: Anti–neurofascin-155 IgG4 antibodies prevent paranodal complex formation in vivo

    doi: 10.1172/JCI124694

    Figure Lengend Snippet: Schematic representation of the pathogenic mechanisms of anti–paranodal protein autoantibodies. ( A ) Representation of a mature node of Ranvier and myelinated axon. Myelin (yellow) covers the axon except at the node of Ranvier (red). At paranodal junctions (green), Nfasc155 interacts with its axonal partners CASPR1/CNTN1. Nfasc155 is also found at Schmidt-Lanterman incisures (dashed green lines). The Schwann cell nucleus is shown in gray. ( B ) In neonatal animals, the progressive enwrapping of axons by Schwann cells induces the formation of paranodal regions at node borders. The injection of anti-Nfasc155 IgG4 (blue) during the neonatal period does not affect myelination or node/paranode formation, but induces the depletion of Nfasc155, and thereby alters the formation of paranodal septate-like junctions. ( C ) At adult age, evidence suggests that the Nfasc155/CASPR1/CNTN1 complex is constantly renewed at paranodes possibly through degradation and replenishment mechanisms. The chronic infusion of anti-Nfasc155 IgG4 (blue) may preclude the regeneration of the paranodal axoglial junction by inducing Nfasc155 depletion, and thereby alter paranode structure and conduction.

    Article Snippet: Purified antibody samples were denatured in SDS sample buffer for 2 minutes at 92°C, loaded on 7.5% SDS-PAGE gels, transferred, and immunoblotted with mouse monoclonal antibodies against human IgG1 (1:1000; 4E3, Abcam), IgG2 (1:1000; HP6014, Abcam), IgG3 (1:1000; HP-6050, Sigma-Aldrich), or IgG4 (1:5000; HP6035, Abcam).

    Techniques: Injection

    Anti-Nfasc155 autoantibodies target surface Schwann cell antigens. ( A ) Sciatic nerve fibers were incubated in vitro with purified anti-Nfasc155 IgG4 from patient CIDP1 for 3 hours, and immunolabeled for IgG4 (green) and CNTN1 (red). ( B – E ) Sciatic nerves were fixed 1 day ( B and D ) or 3 days ( C and E ) after intraneural injections of anti-Nfasc155 IgG4, and immunolabeled for IgG4 (green) and β-catenin (red; B and C ) or CNTN1 (red; D and E ) and Na v channels (blue; D and E ). Note that anti-Nfasc155 IgG4 bound to the surface of the Schwann cells and deposited at the vicinity of the node of Ranvier (double arrowheads) and at adherens junctions along the internode stained here with β-catenin (arrows). However, no penetration across the paranodal region was observed (images are representative of n = 3 independent experiments). Scale bars: 10 μm.

    Journal: The Journal of Clinical Investigation

    Article Title: Anti–neurofascin-155 IgG4 antibodies prevent paranodal complex formation in vivo

    doi: 10.1172/JCI124694

    Figure Lengend Snippet: Anti-Nfasc155 autoantibodies target surface Schwann cell antigens. ( A ) Sciatic nerve fibers were incubated in vitro with purified anti-Nfasc155 IgG4 from patient CIDP1 for 3 hours, and immunolabeled for IgG4 (green) and CNTN1 (red). ( B – E ) Sciatic nerves were fixed 1 day ( B and D ) or 3 days ( C and E ) after intraneural injections of anti-Nfasc155 IgG4, and immunolabeled for IgG4 (green) and β-catenin (red; B and C ) or CNTN1 (red; D and E ) and Na v channels (blue; D and E ). Note that anti-Nfasc155 IgG4 bound to the surface of the Schwann cells and deposited at the vicinity of the node of Ranvier (double arrowheads) and at adherens junctions along the internode stained here with β-catenin (arrows). However, no penetration across the paranodal region was observed (images are representative of n = 3 independent experiments). Scale bars: 10 μm.

    Article Snippet: Purified antibody samples were denatured in SDS sample buffer for 2 minutes at 92°C, loaded on 7.5% SDS-PAGE gels, transferred, and immunoblotted with mouse monoclonal antibodies against human IgG1 (1:1000; 4E3, Abcam), IgG2 (1:1000; HP6014, Abcam), IgG3 (1:1000; HP-6050, Sigma-Aldrich), or IgG4 (1:5000; HP6035, Abcam).

    Techniques: Incubation, In Vitro, Purification, Immunolabeling, Staining

    Passive transfer of anti-Nfasc155 IgG4 affects the formation of paranodal axoglial unit during development. ( A – D ) Newborn rat pups received an i.p. injection of 250 μg of control IgG4 ( A ) or anti-Nfasc155 IgG4 from patient CIDP1 ( B ) on the day of birth ( n = 4 animals for each condition and age). Sciatic nerve fibers were fixed and immunolabeled for voltage-gated sodium channels (Na v ; green) and CASPR1 (red) at postnatal days 0 (P0), 2, 4, and 6. The percentages of Na v clusters with 1 or 2 flanking CASPR1-positive paranodes (double arrowheads) or without CASPR1-positive paranodes (arrowheads) were quantified at each age ( C ), as well as the paranodal length ( D ) ( n = 200–300 nodes or paranodes for each condition and age). Injection of anti-Nfasc155 IgG4 importantly delayed the formation of CASPR1-positive paranodes, and a significantly higher percentage of heminodes without flanking paranodes was observed at P2 (** P

    Journal: The Journal of Clinical Investigation

    Article Title: Anti–neurofascin-155 IgG4 antibodies prevent paranodal complex formation in vivo

    doi: 10.1172/JCI124694

    Figure Lengend Snippet: Passive transfer of anti-Nfasc155 IgG4 affects the formation of paranodal axoglial unit during development. ( A – D ) Newborn rat pups received an i.p. injection of 250 μg of control IgG4 ( A ) or anti-Nfasc155 IgG4 from patient CIDP1 ( B ) on the day of birth ( n = 4 animals for each condition and age). Sciatic nerve fibers were fixed and immunolabeled for voltage-gated sodium channels (Na v ; green) and CASPR1 (red) at postnatal days 0 (P0), 2, 4, and 6. The percentages of Na v clusters with 1 or 2 flanking CASPR1-positive paranodes (double arrowheads) or without CASPR1-positive paranodes (arrowheads) were quantified at each age ( C ), as well as the paranodal length ( D ) ( n = 200–300 nodes or paranodes for each condition and age). Injection of anti-Nfasc155 IgG4 importantly delayed the formation of CASPR1-positive paranodes, and a significantly higher percentage of heminodes without flanking paranodes was observed at P2 (** P

    Article Snippet: Purified antibody samples were denatured in SDS sample buffer for 2 minutes at 92°C, loaded on 7.5% SDS-PAGE gels, transferred, and immunoblotted with mouse monoclonal antibodies against human IgG1 (1:1000; 4E3, Abcam), IgG2 (1:1000; HP6014, Abcam), IgG3 (1:1000; HP-6050, Sigma-Aldrich), or IgG4 (1:5000; HP6035, Abcam).

    Techniques: Injection, Immunolabeling

    Anti-Nfasc155 IgG4 deposits at proximity of nodes in neonatal animals. ( A – C ) Newborn rat pups received daily i.p. injection of 250 μg of control IgG4 ( A ) or anti-Nfasc155 IgG4 from patient CIDP1 ( B ) from the day of birth to P6 (images are representative of n = 3 independent experiments for each age). Sciatic nerve fibers were immunolabeled for voltage-gated sodium channels (Na v ; blue), CASPR1 (green), and IgG (red) at P2 ( A ), P4 ( B ), and P6 ( C ). At P2 and P4, IgG deposits were mostly detected at the borders of Na v clusters, even in Na v clusters lacking CASPR1-positive paranodes (arrowheads). No colocalization with CASPR1 was found even in Na v clusters flanked by 1 or 2 paranodes (double arrowheads). At P6, IgG deposits appeared fainter and were mostly detected at the node vicinity. A weak IgG labeling can be observed at paranodes in some fibers. Isolated node labeling is shown at a higher magnification in the insets. Scale bars: 10 μm.

    Journal: The Journal of Clinical Investigation

    Article Title: Anti–neurofascin-155 IgG4 antibodies prevent paranodal complex formation in vivo

    doi: 10.1172/JCI124694

    Figure Lengend Snippet: Anti-Nfasc155 IgG4 deposits at proximity of nodes in neonatal animals. ( A – C ) Newborn rat pups received daily i.p. injection of 250 μg of control IgG4 ( A ) or anti-Nfasc155 IgG4 from patient CIDP1 ( B ) from the day of birth to P6 (images are representative of n = 3 independent experiments for each age). Sciatic nerve fibers were immunolabeled for voltage-gated sodium channels (Na v ; blue), CASPR1 (green), and IgG (red) at P2 ( A ), P4 ( B ), and P6 ( C ). At P2 and P4, IgG deposits were mostly detected at the borders of Na v clusters, even in Na v clusters lacking CASPR1-positive paranodes (arrowheads). No colocalization with CASPR1 was found even in Na v clusters flanked by 1 or 2 paranodes (double arrowheads). At P6, IgG deposits appeared fainter and were mostly detected at the node vicinity. A weak IgG labeling can be observed at paranodes in some fibers. Isolated node labeling is shown at a higher magnification in the insets. Scale bars: 10 μm.

    Article Snippet: Purified antibody samples were denatured in SDS sample buffer for 2 minutes at 92°C, loaded on 7.5% SDS-PAGE gels, transferred, and immunoblotted with mouse monoclonal antibodies against human IgG1 (1:1000; 4E3, Abcam), IgG2 (1:1000; HP6014, Abcam), IgG3 (1:1000; HP-6050, Sigma-Aldrich), or IgG4 (1:5000; HP6035, Abcam).

    Techniques: Injection, Immunolabeling, Labeling, Isolation

    Anti-Nfasc155 IgG4 disrupts paranodal specialization in ventral spinal nerve. ( A – F ) Teased fibers from L6 ventral roots of animals treated with control IgG4 ( n = 15; A , C , and E ) or anti-Nfasc155 IgG4 from patient CIDP1 ( n = 15; B , D , and F ) were stained for Na v channels (green), CNTN1 (red), and CASPR1 (blue, A and B ), ankyrin-G (Ank-G; blue, C and D ), or Nfasc186 (blue, E and F ). Most nodes of Ranvier were positive for Na v channel, ankyrin-G, and Nfasc186 clusters and were bordered by paranodes stained for CASPR1 and CNTN1 in control animals. By contrast, many nodes lacked CASPR1 or CNTN1 staining at paranodes (arrowheads) in animals chronically injected with anti-Nfasc155 IgG4. Insets in B and D show representative affected nodes. Scale bars: 10 μm.

    Journal: The Journal of Clinical Investigation

    Article Title: Anti–neurofascin-155 IgG4 antibodies prevent paranodal complex formation in vivo

    doi: 10.1172/JCI124694

    Figure Lengend Snippet: Anti-Nfasc155 IgG4 disrupts paranodal specialization in ventral spinal nerve. ( A – F ) Teased fibers from L6 ventral roots of animals treated with control IgG4 ( n = 15; A , C , and E ) or anti-Nfasc155 IgG4 from patient CIDP1 ( n = 15; B , D , and F ) were stained for Na v channels (green), CNTN1 (red), and CASPR1 (blue, A and B ), ankyrin-G (Ank-G; blue, C and D ), or Nfasc186 (blue, E and F ). Most nodes of Ranvier were positive for Na v channel, ankyrin-G, and Nfasc186 clusters and were bordered by paranodes stained for CASPR1 and CNTN1 in control animals. By contrast, many nodes lacked CASPR1 or CNTN1 staining at paranodes (arrowheads) in animals chronically injected with anti-Nfasc155 IgG4. Insets in B and D show representative affected nodes. Scale bars: 10 μm.

    Article Snippet: Purified antibody samples were denatured in SDS sample buffer for 2 minutes at 92°C, loaded on 7.5% SDS-PAGE gels, transferred, and immunoblotted with mouse monoclonal antibodies against human IgG1 (1:1000; 4E3, Abcam), IgG2 (1:1000; HP6014, Abcam), IgG3 (1:1000; HP-6050, Sigma-Aldrich), or IgG4 (1:5000; HP6035, Abcam).

    Techniques: Staining, Injection

    Antibodies to Nfasc155 do not alter the interaction between Nfasc155 and its axonal partners CNTN1 and CASPR1. ( A – C ) For aggregation assays, HEK293 cells were cotransfected with mCherry-tagged Nfasc155 or with GFP-tagged CASPR1 and CNTN1. Cells were then incubated under gentle agitation for 2 hours in the presence of 10 μg of control IgG4, anti-CNTN1 IgG4, or anti-Nfasc155 IgG4 from 3 patients (CIDP1–3). As negative controls, Nfasc155-expressing HEK293 cells were incubated with cells expressing GFP alone (top left panel). Cells were examined with a fluorescence microscope at ×10 objective. Representative fields are shown in A ( n = 3–4 experiments for each condition). Dashed circles highlight cell aggregates with contacts between red and green cells. The percentage of cell clusters with contacts between green and red cells was quantified ( B ), as well as the relative frequency of green cells per aggregate ( C ) ( n = 3–4 experiments for each condition). CASPR1/CNTN1– and Nfasc155-expressing cells form clusters. Anti-CNTN1 IgG4 significantly prevented the formation of cell aggregates (** P

    Journal: The Journal of Clinical Investigation

    Article Title: Anti–neurofascin-155 IgG4 antibodies prevent paranodal complex formation in vivo

    doi: 10.1172/JCI124694

    Figure Lengend Snippet: Antibodies to Nfasc155 do not alter the interaction between Nfasc155 and its axonal partners CNTN1 and CASPR1. ( A – C ) For aggregation assays, HEK293 cells were cotransfected with mCherry-tagged Nfasc155 or with GFP-tagged CASPR1 and CNTN1. Cells were then incubated under gentle agitation for 2 hours in the presence of 10 μg of control IgG4, anti-CNTN1 IgG4, or anti-Nfasc155 IgG4 from 3 patients (CIDP1–3). As negative controls, Nfasc155-expressing HEK293 cells were incubated with cells expressing GFP alone (top left panel). Cells were examined with a fluorescence microscope at ×10 objective. Representative fields are shown in A ( n = 3–4 experiments for each condition). Dashed circles highlight cell aggregates with contacts between red and green cells. The percentage of cell clusters with contacts between green and red cells was quantified ( B ), as well as the relative frequency of green cells per aggregate ( C ) ( n = 3–4 experiments for each condition). CASPR1/CNTN1– and Nfasc155-expressing cells form clusters. Anti-CNTN1 IgG4 significantly prevented the formation of cell aggregates (** P

    Article Snippet: Purified antibody samples were denatured in SDS sample buffer for 2 minutes at 92°C, loaded on 7.5% SDS-PAGE gels, transferred, and immunoblotted with mouse monoclonal antibodies against human IgG1 (1:1000; 4E3, Abcam), IgG2 (1:1000; HP6014, Abcam), IgG3 (1:1000; HP-6050, Sigma-Aldrich), or IgG4 (1:5000; HP6035, Abcam).

    Techniques: Incubation, Expressing, Fluorescence, Microscopy

    Allergen-specific IgG, IgG 1 , IgG 4 levels during the two treatment years. (A) Phl p 1-, (B) Phl p 2-, (C) Phl p 5- and (D) Phl p 6-specific IgG (left graphs), IgG 1 and IgG 4 (right graphs) levels were measured by ELISA (Optical density levels OD 405nm , y -axes) at indicated time points ( x -axes). Lines (Placebo: red circles, BM32: blue squares) represent means with standard deviation shown. Symbols for IgG subclasses are indicated in the legends.

    Journal: EBioMedicine

    Article Title: Two years of treatment with the recombinant grass pollen allergy vaccine BM32 induces a continuously increasing allergen-specific IgG4 response

    doi: 10.1016/j.ebiom.2019.11.006

    Figure Lengend Snippet: Allergen-specific IgG, IgG 1 , IgG 4 levels during the two treatment years. (A) Phl p 1-, (B) Phl p 2-, (C) Phl p 5- and (D) Phl p 6-specific IgG (left graphs), IgG 1 and IgG 4 (right graphs) levels were measured by ELISA (Optical density levels OD 405nm , y -axes) at indicated time points ( x -axes). Lines (Placebo: red circles, BM32: blue squares) represent means with standard deviation shown. Symbols for IgG subclasses are indicated in the legends.

    Article Snippet: Subclasses IgG1 and IgG4 were detected using mouse monoclonal anti-human IgG1 (clone JDC-1) and IgG4 (clone JDC-14) antibodies (1:1000, BD Bioscience, San Jose, CA, USA) followed by HRP-coupled sheep anti-mouse IgG (1:2000) (GE Healthcare, Waukesha, WI, USA).

    Techniques: Enzyme-linked Immunosorbent Assay, Standard Deviation

    Serum IgG1 levels of 13 children with TS before, during, and after an exacerbation of tics. Patients showed increased IgG1 levels during exacerbation: F(2, 24) = 2.63, p = .09.

    Journal: Brain, behavior, and immunity

    Article Title: ALTERED IMMUNOGLOBULIN PROFILES IN CHILDREN WITH TOURETTE SYNDROME

    doi: 10.1016/j.bbi.2010.12.003

    Figure Lengend Snippet: Serum IgG1 levels of 13 children with TS before, during, and after an exacerbation of tics. Patients showed increased IgG1 levels during exacerbation: F(2, 24) = 2.63, p = .09.

    Article Snippet: Ig concentrations were measured in archived sera using a Luminex based Ig Isotyping reagent kit for measurement of IgG1, IgG2, IgG3, and IgG4, and IgM, IgA, and IgE (Bio-Rad Laboratories, Hercules, CA, catalog number 171-A3001M) in 10,000 fold diluted serum samples according to manufacturer’s instructions.

    Techniques:

    Peanut specific (A) IgE, (B) IgG4, (C) IgE/IgG4 ratio and (D) skin prick test results at baseline and after a year of therapy for participants with proven peanut allergy (* indicates p

    Journal: Allergy, Asthma, and Clinical Immunology : Official Journal of the Canadian Society of Allergy and Clinical Immunology

    Article Title: Phase 1 results of safety and tolerability in a rush oral immunotherapy protocol to multiple foods using Omalizumab

    doi: 10.1186/1710-1492-10-7

    Figure Lengend Snippet: Peanut specific (A) IgE, (B) IgG4, (C) IgE/IgG4 ratio and (D) skin prick test results at baseline and after a year of therapy for participants with proven peanut allergy (* indicates p

    Article Snippet: Sera were analyzed for peanut-specific IgE and IgG4 levels at John Hopkins Allergy and Clinical Immunology Reference Laboratory by immunoCAP FEIA (Thermofisher Scientific/Phadia, Kalmazoo, MI).

    Techniques:

    Analysis of neutralization sensitivity of Env variants taken from donor 45 at three time points to autologous serum IgG (left panels) and autologous CD4bs MAbs VRC01, VRC03, VRC06, and VRC06b (right panels). The horizontal bars indicate the group geometric

    Journal: Journal of Virology

    Article Title: Selection Pressure on HIV-1 Envelope by Broadly Neutralizing Antibodies to the Conserved CD4-Binding Site

    doi: 10.1128/JVI.07139-11

    Figure Lengend Snippet: Analysis of neutralization sensitivity of Env variants taken from donor 45 at three time points to autologous serum IgG (left panels) and autologous CD4bs MAbs VRC01, VRC03, VRC06, and VRC06b (right panels). The horizontal bars indicate the group geometric

    Article Snippet: The concentrations of purified serum IgG or IgG MAbs were determined using NanoDrop (Thermo Scientific) with 1.40 as the extinction coefficient.

    Techniques: Neutralization

    Comparative analysis of different serodiagnostic assays from serum samples obtained from PKDL patients, Healthy controls (EC and NEC) and disease control groups (Leprosy and Vitiligo). (A) Comparative evaluation of PEG CIC at 450 nm obtained from PEG precipitation of PKDL patients’ sera; cut-off 0.274. (B) Comparative evaluation of ELISA reactivity of anti-leishmanial antibody (IgG) in serum against Leishmania antigen; cut-off 0.350. (C) Comparative evaluation of ELISA reactivity of anti-leishmanial antibody (IgM) in serum against Leishmania antigen; cut-off 0.191. (D) Comparative evaluation of Glyco CIC assay based on CICs isolated from sera of PKDL patients; cut-off 0.091. The study population in all cases comprised of panel of PKDL patients (PKDL; n = 90), Endemic Healthy controls (n = 19), Non-Endemic Healthy controls (n = 34) and other disease (OD; n = 83) including Leprosy (n = 37) and Vitiligo (n = 46). Each sample was tested in triplicates and mean was taken. Each dot represent mean of single sample and the black dotted horizontal lines represent cut-off values. The cut-off value of the assays was established as means + 3 SD of 136 controls. Significance was determined by unpaired Student's t-test at 95% confidence intervals and p-values

    Journal: PLoS ONE

    Article Title: Glycoproteins in circulating immune complexes are biomarkers of patients with Indian PKDL: A study from endemic districts of West Bengal, India

    doi: 10.1371/journal.pone.0192302

    Figure Lengend Snippet: Comparative analysis of different serodiagnostic assays from serum samples obtained from PKDL patients, Healthy controls (EC and NEC) and disease control groups (Leprosy and Vitiligo). (A) Comparative evaluation of PEG CIC at 450 nm obtained from PEG precipitation of PKDL patients’ sera; cut-off 0.274. (B) Comparative evaluation of ELISA reactivity of anti-leishmanial antibody (IgG) in serum against Leishmania antigen; cut-off 0.350. (C) Comparative evaluation of ELISA reactivity of anti-leishmanial antibody (IgM) in serum against Leishmania antigen; cut-off 0.191. (D) Comparative evaluation of Glyco CIC assay based on CICs isolated from sera of PKDL patients; cut-off 0.091. The study population in all cases comprised of panel of PKDL patients (PKDL; n = 90), Endemic Healthy controls (n = 19), Non-Endemic Healthy controls (n = 34) and other disease (OD; n = 83) including Leprosy (n = 37) and Vitiligo (n = 46). Each sample was tested in triplicates and mean was taken. Each dot represent mean of single sample and the black dotted horizontal lines represent cut-off values. The cut-off value of the assays was established as means + 3 SD of 136 controls. Significance was determined by unpaired Student's t-test at 95% confidence intervals and p-values

    Article Snippet: The wells were then incubated for 1hr at RT with HRP-conjugated anti-human monoclonal IgG (1:15,000) (Sigma-Aldrich, Cat # :A0170), monoclonal IgG1 (1:1000) (Thermo Fisher Scientific, Cat#: A-10648, USA), monoclonal IgG2 (1:1000) (Thermo Fisher Scientific, Cat#:MH1772, USA), monoclonal IgG3 (1:1000) (Thermo Fisher Scientific, Cat#:05–3620,USA), monoclonal IgG4 (1:1000) (Thermo Fisher Scientific, Cat#:A-10654,USA), and polyclonal IgM (1:10,000) (Sigma-Aldrich, Cat#:A6907), respectively, to measure the levels of IgM, IgG and IgG subclasses of anti-leishmanial antibodies with Tetramethylbenzidine (TMB).

    Techniques: Enzyme-linked Immunosorbent Assay, Isolation

    IgG, IgG1 and IgG2a antibody responses against total mixed and individual Brucella L7/L12, Omp16, Omp19 and SOD proteins in pregnant sheep (A) and goats (B) at 0, 21, 42, 63 days post-vaccination (DPV) by ELISA. Pregnant sheep and goats in the group I (Flu-BA_Omp19-SOD) were immunized twice concurrently via the subcutaneous and conjunctival routes at an interval of 21 days with vaccines generated from the influenza viral vectors (IVV) subtypes H5N1 (prime vaccination) and H1N1 (booster vaccination). The vaccination of animals of group II (Flu-BA_Omp19-SOD_TV) was carried out in the same way as in group I, but only the vaccine generated from IVV subtypes H5N1 was used, which was administered three times at 21 days intervals. Sheep and goats in the negative control group (IV) were vaccinated with 20% Montanide Gel01 adjuvant in PBS three times at 21 days intervals. Data are presented as optical density (OD) ± standard deviations; * P = 0 . 04 to P

    Journal: PLoS ONE

    Article Title: Improved influenza viral vector based Brucella abortus vaccine induces robust B and T-cell responses and protection against Brucella melitensis infection in pregnant sheep and goats

    doi: 10.1371/journal.pone.0186484

    Figure Lengend Snippet: IgG, IgG1 and IgG2a antibody responses against total mixed and individual Brucella L7/L12, Omp16, Omp19 and SOD proteins in pregnant sheep (A) and goats (B) at 0, 21, 42, 63 days post-vaccination (DPV) by ELISA. Pregnant sheep and goats in the group I (Flu-BA_Omp19-SOD) were immunized twice concurrently via the subcutaneous and conjunctival routes at an interval of 21 days with vaccines generated from the influenza viral vectors (IVV) subtypes H5N1 (prime vaccination) and H1N1 (booster vaccination). The vaccination of animals of group II (Flu-BA_Omp19-SOD_TV) was carried out in the same way as in group I, but only the vaccine generated from IVV subtypes H5N1 was used, which was administered three times at 21 days intervals. Sheep and goats in the negative control group (IV) were vaccinated with 20% Montanide Gel01 adjuvant in PBS three times at 21 days intervals. Data are presented as optical density (OD) ± standard deviations; * P = 0 . 04 to P

    Article Snippet: A donkey anti-ruminant IgG horseradish peroxidase conjugate (Sigma, St. Louis, MO, USA) and monoclonal antibodies specific for sheep IgG1 and IgG2 (Novus Biologicals, Littleton, CO, USA) were used for detection.

    Techniques: Enzyme-linked Immunosorbent Assay, Generated, Negative Control

    Anti-Dsg3 autoantibodies detected in healthy blood donors do not lead to Dsg3 internalization in vitro . HaCaT cells were incubated with IgG from healthy individuals (NH-IgG), IgG from a pemphigus vulgaris patient (PV-IgG) or with Dsg3-positive plasma samples from healthy blood donors (samples #1382, #3167 and #9607), as well as with a Dsg3-negative sample (sample #4025). For all experimental conditions ( n = 3), the IgG concentration was 1 mg/ml (with the exception of the diluted PV-IgG samples). While NH-IgG and 4025 did not induce Dsg3 internalization, PV-IgG at 1:1 and 1:100 dilutions led to Dsg3 internalization. At low dilutions, PV-IgG and all tested Dsg3-positive samples from healthy blood donors did not show any effects on Dsg3 internalization

    Journal: Orphanet Journal of Rare Diseases

    Article Title: Prevalence of pemphigus and pemphigoid autoantibodies in the general population

    doi: 10.1186/s13023-015-0278-x

    Figure Lengend Snippet: Anti-Dsg3 autoantibodies detected in healthy blood donors do not lead to Dsg3 internalization in vitro . HaCaT cells were incubated with IgG from healthy individuals (NH-IgG), IgG from a pemphigus vulgaris patient (PV-IgG) or with Dsg3-positive plasma samples from healthy blood donors (samples #1382, #3167 and #9607), as well as with a Dsg3-negative sample (sample #4025). For all experimental conditions ( n = 3), the IgG concentration was 1 mg/ml (with the exception of the diluted PV-IgG samples). While NH-IgG and 4025 did not induce Dsg3 internalization, PV-IgG at 1:1 and 1:100 dilutions led to Dsg3 internalization. At low dilutions, PV-IgG and all tested Dsg3-positive samples from healthy blood donors did not show any effects on Dsg3 internalization

    Article Snippet: After incubation with human plasma, ELISA plates were incubated with biotin-conjugated mouse anti-human IgG1, IgG2, IgG3 and IgG4 antibodies (Invitrogen, Camarillo, CA), followed by incubation with peroxidase-conjugated donkey anti-mouse IgG antibody (minimal cross-reaction to serum proteins of other species, Jackson ImmunoResearch Laboratories, West Grove, PA).

    Techniques: In Vitro, Incubation, Concentration Assay

    Anti-BP180-NC16A autoantibodies from healthy individuals bind to different epitopes within the NC16A domain. Samples from the indicated groups were tested for their IgG reactivity against NC16A2-3 (RSILPYGDSMDRIEKDRLQGMAPAAGADL). As reported previously [ 29 , 30 ], high reactivity with this epitope was observed in BP patients. By contrast, only a minority of samples from healthy individuals with IgG reactivity against NC16A showed reactivity with this epitope. Abbreviations: NHS: normal human sera

    Journal: Orphanet Journal of Rare Diseases

    Article Title: Prevalence of pemphigus and pemphigoid autoantibodies in the general population

    doi: 10.1186/s13023-015-0278-x

    Figure Lengend Snippet: Anti-BP180-NC16A autoantibodies from healthy individuals bind to different epitopes within the NC16A domain. Samples from the indicated groups were tested for their IgG reactivity against NC16A2-3 (RSILPYGDSMDRIEKDRLQGMAPAAGADL). As reported previously [ 29 , 30 ], high reactivity with this epitope was observed in BP patients. By contrast, only a minority of samples from healthy individuals with IgG reactivity against NC16A showed reactivity with this epitope. Abbreviations: NHS: normal human sera

    Article Snippet: After incubation with human plasma, ELISA plates were incubated with biotin-conjugated mouse anti-human IgG1, IgG2, IgG3 and IgG4 antibodies (Invitrogen, Camarillo, CA), followed by incubation with peroxidase-conjugated donkey anti-mouse IgG antibody (minimal cross-reaction to serum proteins of other species, Jackson ImmunoResearch Laboratories, West Grove, PA).

    Techniques:

    Serial dilution tests performed using PROTIA™ Specific IgG4®. Patient sera were diluted with immunoglobulin-depleted serum. N * : negative. Specific IgG4

    Journal: Yonsei Medical Journal

    Article Title: Validation of a Multiplex Assay for Measuring Specific IgG4

    doi: 10.3349/ymj.2020.61.6.524

    Figure Lengend Snippet: Serial dilution tests performed using PROTIA™ Specific IgG4®. Patient sera were diluted with immunoglobulin-depleted serum. N * : negative. Specific IgG4

    Article Snippet: PROTIA™ Specific IgG4® may be applicable to patients with Crohn's disease, for whom dietary restrictions are recommended.

    Techniques: Serial Dilution

    Measured sIgG4 values for 12 allergens that were positive in both ImmunoCAP® and PROTIA™ Specific IgG4®.

    Journal: Yonsei Medical Journal

    Article Title: Validation of a Multiplex Assay for Measuring Specific IgG4

    doi: 10.3349/ymj.2020.61.6.524

    Figure Lengend Snippet: Measured sIgG4 values for 12 allergens that were positive in both ImmunoCAP® and PROTIA™ Specific IgG4®.

    Article Snippet: PROTIA™ Specific IgG4® may be applicable to patients with Crohn's disease, for whom dietary restrictions are recommended.

    Techniques:

    Infectious mastoiditis mimicking IgG4 related mastoiditis. The storiform type fibrosis is more apparent in case 4 ( a ) than case 5 ( c ). However, both cases show markedly increased numbers of IgG4 positive plasma cells ( b , d )

    Journal: Head and Neck Pathology

    Article Title: Recurrent Mastoiditis Mimics IgG4 Related Disease: A Potential Diagnostic Pitfall

    doi: 10.1007/s12105-016-0710-0

    Figure Lengend Snippet: Infectious mastoiditis mimicking IgG4 related mastoiditis. The storiform type fibrosis is more apparent in case 4 ( a ) than case 5 ( c ). However, both cases show markedly increased numbers of IgG4 positive plasma cells ( b , d )

    Article Snippet: Th2 cytokine responses, particularly those driven by interleukin-4 and interleukin-10, mediate B cell class switching to IgG4 [ ].

    Techniques:

    Case 1: IgG4 related mastoiditis. Dense lymphoplasmacytic infiltrate with storiform type fibrosis ( a , b ). The infiltrate involves the bone ( c ). Arrow indicates lamellar bone and the periosteum is marked with an asterisk . An immunohistochemical stain for IgG4 shows elevated numbers of IgG4 positive plasma cells ( d )

    Journal: Head and Neck Pathology

    Article Title: Recurrent Mastoiditis Mimics IgG4 Related Disease: A Potential Diagnostic Pitfall

    doi: 10.1007/s12105-016-0710-0

    Figure Lengend Snippet: Case 1: IgG4 related mastoiditis. Dense lymphoplasmacytic infiltrate with storiform type fibrosis ( a , b ). The infiltrate involves the bone ( c ). Arrow indicates lamellar bone and the periosteum is marked with an asterisk . An immunohistochemical stain for IgG4 shows elevated numbers of IgG4 positive plasma cells ( d )

    Article Snippet: Th2 cytokine responses, particularly those driven by interleukin-4 and interleukin-10, mediate B cell class switching to IgG4 [ ].

    Techniques: Immunohistochemistry, Staining

    Whole-cell ELISA antibody levels against strain H44/76 in children with meningococcal infection, controls, siblings of children with meningococcal infection, and adults in relation to age. Means of the antibody concentration and standard errors of the means are reported for total IgG (a), IgG1 (b), IgG2 (c), IgG3 (d), IgG4 (e), and IgM (f).

    Journal: Infection and Immunity

    Article Title: Humoral Immune Responses to Neisseria meningitidis in Children

    doi:

    Figure Lengend Snippet: Whole-cell ELISA antibody levels against strain H44/76 in children with meningococcal infection, controls, siblings of children with meningococcal infection, and adults in relation to age. Means of the antibody concentration and standard errors of the means are reported for total IgG (a), IgG1 (b), IgG2 (c), IgG3 (d), IgG4 (e), and IgM (f).

    Article Snippet: Conversely, IgG2 (93 ± 31 U) (Fig. c), IgG4 (202 ± 62 U) (Fig. e), and IgM (421 ± 72 U) (Fig. f) were not significantly raised at 9 weeks (median) following natural infection compared with controls (112 ± 20, 155 ± 41, and 476 ± 58 U, respectively).

    Techniques: Enzyme-linked Immunosorbent Assay, Infection, Concentration Assay

    HIV-specific responses and immune response capability of the case at 9.5 years of age. a Detection of HIV-specific antibodies at 9.5 years of age by western blot. The case antibody profile is compared with controls that are a high positive, low positive and HIV-negative. HIV proteins corresponding to bands in the blots are shown in the left grey-shaded block; the case profile was positive for the core proteins indicated in pink. b Quantitation of HIV-specific antibodies by multiplex bead array for all isotypes and subclasses (indicated on the left side—IgG1, IgG2, IgG3, IgG4, IgM, IgA1, IgA2) against gp41, Gag, RT, gp120, Nef, Tat, Vpu, peptide Vpu9 and V1V2 scaffold antigens (indicated at the top). Results are expressed as mean fluorescence intensities (MFI)), the colour key shows ranges of MFI according to colour intensity (the darker the more HIV-specific antibody detected). A result is considered positive if above the cut-off (mean ± 3 SD) determined from eight adult uninfected controls. Vpu9 amino acid sequence: STMVDMGHLRLLDVNDL. c Proportions of natural killer (NK) cells that respond to anti-CD16 antibody, and CD4+ and CD8+ T cells that respond to staphylococcal enterotoxin B (SEB) in a whole blood intracellular cytokine (ICC) assay that measures induction of interferon-γ (IFN-γ) and interleukin-2 (IL-2). HIV-uninfected adult reference values for comparison ( n = 21; median % and range)—natural killer (NK) anti-CD16%: 37.92 (12–67.6), CD4 SEB%: 6.04 (0.25–11.91), CD8 SEB%: 5.82 (0.18–18.94). d A weak positive CD4+ T cell response to Gag (0.116%) in the absence of a detectable CD8+ T cell response to Gag (

    Journal: Nature Communications

    Article Title: A child with perinatal HIV infection and long-term sustained virological control following antiretroviral treatment cessation

    doi: 10.1038/s41467-019-08311-0

    Figure Lengend Snippet: HIV-specific responses and immune response capability of the case at 9.5 years of age. a Detection of HIV-specific antibodies at 9.5 years of age by western blot. The case antibody profile is compared with controls that are a high positive, low positive and HIV-negative. HIV proteins corresponding to bands in the blots are shown in the left grey-shaded block; the case profile was positive for the core proteins indicated in pink. b Quantitation of HIV-specific antibodies by multiplex bead array for all isotypes and subclasses (indicated on the left side—IgG1, IgG2, IgG3, IgG4, IgM, IgA1, IgA2) against gp41, Gag, RT, gp120, Nef, Tat, Vpu, peptide Vpu9 and V1V2 scaffold antigens (indicated at the top). Results are expressed as mean fluorescence intensities (MFI)), the colour key shows ranges of MFI according to colour intensity (the darker the more HIV-specific antibody detected). A result is considered positive if above the cut-off (mean ± 3 SD) determined from eight adult uninfected controls. Vpu9 amino acid sequence: STMVDMGHLRLLDVNDL. c Proportions of natural killer (NK) cells that respond to anti-CD16 antibody, and CD4+ and CD8+ T cells that respond to staphylococcal enterotoxin B (SEB) in a whole blood intracellular cytokine (ICC) assay that measures induction of interferon-γ (IFN-γ) and interleukin-2 (IL-2). HIV-uninfected adult reference values for comparison ( n = 21; median % and range)—natural killer (NK) anti-CD16%: 37.92 (12–67.6), CD4 SEB%: 6.04 (0.25–11.91), CD8 SEB%: 5.82 (0.18–18.94). d A weak positive CD4+ T cell response to Gag (0.116%) in the absence of a detectable CD8+ T cell response to Gag (

    Article Snippet: HIV-specific antibody isotypes were detected by adding 50 µL per well at 2 µg mL−1 , R -phycoerthyrin-conjugated mouse anti-human IgG1 to IgG4 (Cat. Nos 9052-09, 9070-09, 9210-09, 9200-09, respectively, Southern Biotech, USA), mouse anti-human IgM (Cat. No. 9020-09, Southern Biotech), mouse anti-human IgA1 (Cat. No. 9130-09, Southern Biotech) or mouse anti-human IgA2 (Cat. No. 9140-09, Southern Biotech) with shaking (in the dark at room temperature) followed by three washes.

    Techniques: Western Blot, Blocking Assay, Quantitation Assay, Multiplex Assay, Fluorescence, Sequencing, Immunocytochemistry

    A strong correlation between autoantibody binding to IgG complexed with each HLA-DR allele and the odds ratio for that allele’s association with RA. ( A ) The IgGH was cotransfected with Ig light chain (L), Ii, and HLA-DRα in combination

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: Autoantibodies to IgG/HLA class II complexes are associated with rheumatoid arthritis susceptibility

    doi: 10.1073/pnas.1401105111

    Figure Lengend Snippet: A strong correlation between autoantibody binding to IgG complexed with each HLA-DR allele and the odds ratio for that allele’s association with RA. ( A ) The IgGH was cotransfected with Ig light chain (L), Ii, and HLA-DRα in combination

    Article Snippet: The membranes were incubated with anti–HLA-DRα Ab (Santa Cruz Biotechnology), followed by anti-rabbit IgG (Thermo Fisher Scientific) or anti-human IgG (Jackson ImmunoResearch) Ab.

    Techniques: Binding Assay

    IgGH is transported to the cell surface by HLA-DR. ( A ) The secreted forms of IgGHs cloned from human PBMCs that have different V regions were cotransfected with HLA-DRα, DRB1*04:04 (HLA-DR4), and GFP. Cell surface IgG on GFP-expressing cells was

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: Autoantibodies to IgG/HLA class II complexes are associated with rheumatoid arthritis susceptibility

    doi: 10.1073/pnas.1401105111

    Figure Lengend Snippet: IgGH is transported to the cell surface by HLA-DR. ( A ) The secreted forms of IgGHs cloned from human PBMCs that have different V regions were cotransfected with HLA-DRα, DRB1*04:04 (HLA-DR4), and GFP. Cell surface IgG on GFP-expressing cells was

    Article Snippet: The membranes were incubated with anti–HLA-DRα Ab (Santa Cruz Biotechnology), followed by anti-rabbit IgG (Thermo Fisher Scientific) or anti-human IgG (Jackson ImmunoResearch) Ab.

    Techniques: Clone Assay, Expressing

    Correlation of the effusion κ/λ free L chains (FLC) ratio with immunoglobulin (Ig)G4 + plasma cell counts (a) and IgG4 + /IgG + plasma cell ratio (b) in the pleura of patients in the IgG4 + group. *One‐tailed P ‐value.

    Journal: Clinical and Experimental Immunology

    Article Title: Association of immunoglobulin G4 and free light chain with idiopathic pleural effusion

    doi: 10.1111/cei.12999

    Figure Lengend Snippet: Correlation of the effusion κ/λ free L chains (FLC) ratio with immunoglobulin (Ig)G4 + plasma cell counts (a) and IgG4 + /IgG + plasma cell ratio (b) in the pleura of patients in the IgG4 + group. *One‐tailed P ‐value.

    Article Snippet: After transferring the proteins onto PVDF membranes, γ4, κ and λ chains on the blots were probed with biotinylated anti‐IgG4 (Sigma; cat. no. B3648)/horseradish peroxidase‐conjugated Extavidin (Sigma; cat. no. E2886), peroxidase‐conjugated anti‐κ and anti‐λ light chain antibodies (Bio‐Rad; cat. nos STAR127P and STAR129P), respectively.

    Techniques: One-tailed Test

    Receiver operating characteristic (ROC) analysis on diagnostic utility of immunoglobulin (Ig)G4 and κ/λ ratio for distinguishing patients between the IgG4 − and IgG + groups. Cut‐off value for κ/λ ratio, 1·42; sensitivity, 0·87; specificity, 0·83. Area under the curve (AUC), 0·88; 95% confidence interval (CI) for the AUC, 0·74 – 1·00. Cut‐off value for IgG4/IgG ratio, 2·75%; sensitivity, 0·75; specificity, 0·74; AUC, 0·80; 95% CI for the AUC, 0·66–0·94.

    Journal: Clinical and Experimental Immunology

    Article Title: Association of immunoglobulin G4 and free light chain with idiopathic pleural effusion

    doi: 10.1111/cei.12999

    Figure Lengend Snippet: Receiver operating characteristic (ROC) analysis on diagnostic utility of immunoglobulin (Ig)G4 and κ/λ ratio for distinguishing patients between the IgG4 − and IgG + groups. Cut‐off value for κ/λ ratio, 1·42; sensitivity, 0·87; specificity, 0·83. Area under the curve (AUC), 0·88; 95% confidence interval (CI) for the AUC, 0·74 – 1·00. Cut‐off value for IgG4/IgG ratio, 2·75%; sensitivity, 0·75; specificity, 0·74; AUC, 0·80; 95% CI for the AUC, 0·66–0·94.

    Article Snippet: After transferring the proteins onto PVDF membranes, γ4, κ and λ chains on the blots were probed with biotinylated anti‐IgG4 (Sigma; cat. no. B3648)/horseradish peroxidase‐conjugated Extavidin (Sigma; cat. no. E2886), peroxidase‐conjugated anti‐κ and anti‐λ light chain antibodies (Bio‐Rad; cat. nos STAR127P and STAR129P), respectively.

    Techniques: Diagnostic Assay

    Comparison of pleural fluid levels of the κ and λ free L chains (FLC) (a,b) and κ/λ ratio (c). Median and interquartile ranges are shown. (−), Immunoglobulin (Ig)G4 − group; (+), IgG4 + group.

    Journal: Clinical and Experimental Immunology

    Article Title: Association of immunoglobulin G4 and free light chain with idiopathic pleural effusion

    doi: 10.1111/cei.12999

    Figure Lengend Snippet: Comparison of pleural fluid levels of the κ and λ free L chains (FLC) (a,b) and κ/λ ratio (c). Median and interquartile ranges are shown. (−), Immunoglobulin (Ig)G4 − group; (+), IgG4 + group.

    Article Snippet: After transferring the proteins onto PVDF membranes, γ4, κ and λ chains on the blots were probed with biotinylated anti‐IgG4 (Sigma; cat. no. B3648)/horseradish peroxidase‐conjugated Extavidin (Sigma; cat. no. E2886), peroxidase‐conjugated anti‐κ and anti‐λ light chain antibodies (Bio‐Rad; cat. nos STAR127P and STAR129P), respectively.

    Techniques:

    Comparison of pleural fluid levels of immunoglobulins between the immunoglobulin (Ig)G4 − and IgG4 + groups. (−), IgG4 − group; (+), IgG4 + group. Median and interquartile ranges are shown.

    Journal: Clinical and Experimental Immunology

    Article Title: Association of immunoglobulin G4 and free light chain with idiopathic pleural effusion

    doi: 10.1111/cei.12999

    Figure Lengend Snippet: Comparison of pleural fluid levels of immunoglobulins between the immunoglobulin (Ig)G4 − and IgG4 + groups. (−), IgG4 − group; (+), IgG4 + group. Median and interquartile ranges are shown.

    Article Snippet: After transferring the proteins onto PVDF membranes, γ4, κ and λ chains on the blots were probed with biotinylated anti‐IgG4 (Sigma; cat. no. B3648)/horseradish peroxidase‐conjugated Extavidin (Sigma; cat. no. E2886), peroxidase‐conjugated anti‐κ and anti‐λ light chain antibodies (Bio‐Rad; cat. nos STAR127P and STAR129P), respectively.

    Techniques:

    Analysis of the clonality of the effusion immunoglobulin (Ig)G4 antibodies of patients in the IgG4 + group by two‐dimensional electrophoresis (2‐DE). Control IgG4 κ myeloma protein from Sigma (cat no. I4639) (a –c). Effusion IgG4 antibodies of representative cases with abnormal IgG4 levels (d–i). The H and L chains were probed with anti‐IgG4‐Fc (left), anti‐κ chain (middle) and anti‐λ chain (right) antibodies.

    Journal: Clinical and Experimental Immunology

    Article Title: Association of immunoglobulin G4 and free light chain with idiopathic pleural effusion

    doi: 10.1111/cei.12999

    Figure Lengend Snippet: Analysis of the clonality of the effusion immunoglobulin (Ig)G4 antibodies of patients in the IgG4 + group by two‐dimensional electrophoresis (2‐DE). Control IgG4 κ myeloma protein from Sigma (cat no. I4639) (a –c). Effusion IgG4 antibodies of representative cases with abnormal IgG4 levels (d–i). The H and L chains were probed with anti‐IgG4‐Fc (left), anti‐κ chain (middle) and anti‐λ chain (right) antibodies.

    Article Snippet: After transferring the proteins onto PVDF membranes, γ4, κ and λ chains on the blots were probed with biotinylated anti‐IgG4 (Sigma; cat. no. B3648)/horseradish peroxidase‐conjugated Extavidin (Sigma; cat. no. E2886), peroxidase‐conjugated anti‐κ and anti‐λ light chain antibodies (Bio‐Rad; cat. nos STAR127P and STAR129P), respectively.

    Techniques: Electrophoresis

    (A) Fab fragments from FS-1 IgG4 bind wDsg1(blue), bind partially to Dsg1-m1 (red) and do not bind Dsg1-m2 (green) or to wDsg4 (purple) that overlap in a single line. (B) Same Fab fragments from FS1 IgG4 are pathogenic when tested on the mouse model. The animals show skin blisters, which are subcorneal and the Fab fragments are bound to the surface of keratinocytes undergoing detachment at the site of injury by direct immunofluorescence (IF). Orignal magnification of histology and direct IF, x200.

    Journal: Journal of autoimmunity

    Article Title: Pathogenic IgG4 autoantibodies from endemic pemphigus foliaceus recognize a desmoglein-1 conformational epitope

    doi: 10.1016/j.jaut.2017.12.017

    Figure Lengend Snippet: (A) Fab fragments from FS-1 IgG4 bind wDsg1(blue), bind partially to Dsg1-m1 (red) and do not bind Dsg1-m2 (green) or to wDsg4 (purple) that overlap in a single line. (B) Same Fab fragments from FS1 IgG4 are pathogenic when tested on the mouse model. The animals show skin blisters, which are subcorneal and the Fab fragments are bound to the surface of keratinocytes undergoing detachment at the site of injury by direct immunofluorescence (IF). Orignal magnification of histology and direct IF, x200.

    Article Snippet: Commercially available human IgG4 (Sigma Aldrich, St Louis, MO, USA) was injected as a control.

    Techniques: Immunofluorescence

    Identification of key amino acid residues for FS IgG4 binding ( A )The locations of the mutated residues (blue, lime green, yellow and dark purple) in the EC1 epitope (red) are shown on the homology model of the Dsg1/Dsc1 heterodimer (light (Dsg1) and dark (Dsc1) gray). ( B ) Affinity-purified FS1 IgG4 antibodies on the llama anti-human IgG4 and Dsg1-EC1/Dsg3 bb columns were tested against wDsg1 (blue), Dsg1-m1 (red), Dsg1-m2 (green) and wDsg3 (purple) by ELISA. A stock of IgG4 (0.3 mg/ml) from each patient at a dilution of 1:5,000 was tested on microtiter wells coated with different amounts of the recombinant protein (from 12.5 ng/well to 50 ng/well). Peroxidase-labeled mouse anti-human IgG4 at a dilution of 1:2,000 was used as an indicator. The results are expressed in OD 490nm units. ( C ) Affinity-purified FS1 IgG4 antibodies on the llama anti-human IgG4 and Dsg1-EC1/Dsg4 bb columns were tested against wDsg1 (blue), Dsg1-m1 (red), Dsg1-m3 (green) and wDsg4 (purple) by ELISA. A stock of IgG4 (0.3 mg/ml) from each patient at a dilution of 1:5,000 was tested on microtiter wells coated with different amounts of the recombinant protein (from 12.5 ng/well to 50 ng/well). Peroxidase-labeled mouse anti-human IgG4 at a dilution of 1:2,000 was used as an indicator. The results are expressed in OD 490nm units.

    Journal: Journal of autoimmunity

    Article Title: Pathogenic IgG4 autoantibodies from endemic pemphigus foliaceus recognize a desmoglein-1 conformational epitope

    doi: 10.1016/j.jaut.2017.12.017

    Figure Lengend Snippet: Identification of key amino acid residues for FS IgG4 binding ( A )The locations of the mutated residues (blue, lime green, yellow and dark purple) in the EC1 epitope (red) are shown on the homology model of the Dsg1/Dsc1 heterodimer (light (Dsg1) and dark (Dsc1) gray). ( B ) Affinity-purified FS1 IgG4 antibodies on the llama anti-human IgG4 and Dsg1-EC1/Dsg3 bb columns were tested against wDsg1 (blue), Dsg1-m1 (red), Dsg1-m2 (green) and wDsg3 (purple) by ELISA. A stock of IgG4 (0.3 mg/ml) from each patient at a dilution of 1:5,000 was tested on microtiter wells coated with different amounts of the recombinant protein (from 12.5 ng/well to 50 ng/well). Peroxidase-labeled mouse anti-human IgG4 at a dilution of 1:2,000 was used as an indicator. The results are expressed in OD 490nm units. ( C ) Affinity-purified FS1 IgG4 antibodies on the llama anti-human IgG4 and Dsg1-EC1/Dsg4 bb columns were tested against wDsg1 (blue), Dsg1-m1 (red), Dsg1-m3 (green) and wDsg4 (purple) by ELISA. A stock of IgG4 (0.3 mg/ml) from each patient at a dilution of 1:5,000 was tested on microtiter wells coated with different amounts of the recombinant protein (from 12.5 ng/well to 50 ng/well). Peroxidase-labeled mouse anti-human IgG4 at a dilution of 1:2,000 was used as an indicator. The results are expressed in OD 490nm units.

    Article Snippet: Commercially available human IgG4 (Sigma Aldrich, St Louis, MO, USA) was injected as a control.

    Techniques: Binding Assay, Affinity Purification, Enzyme-linked Immunosorbent Assay, Recombinant, Labeling

    Inhibition of desmosomal cadherin-mediated bead aggregation by FS antibodies (A) Co-aggregation of cadherin-coated red and green fluorescent beads mixed in homophilic (Dsg1/Dsg1, Dsg3/Dsg3, Dsc1/Dsc1) or heterophilic combinations (Dsg1/Dsc1 and Dsg3/Dsc1). (B) Inhibition experiments showing heterophilic aggregation of green Dsc1-coated beads with red Dsg1 (top panels) or Dsg3- coated beads (bottom panels) pre-incubated with purified IgG4 fractions from FS patients or from a healthy donor (see methods for details). (C) Dose-dependence of IgG4 Fab mediated inhibition of Dsg1/Dsc1 bead aggregation from donors FS 1 and FS17. Scale bar 0.5mm.

    Journal: Journal of autoimmunity

    Article Title: Pathogenic IgG4 autoantibodies from endemic pemphigus foliaceus recognize a desmoglein-1 conformational epitope

    doi: 10.1016/j.jaut.2017.12.017

    Figure Lengend Snippet: Inhibition of desmosomal cadherin-mediated bead aggregation by FS antibodies (A) Co-aggregation of cadherin-coated red and green fluorescent beads mixed in homophilic (Dsg1/Dsg1, Dsg3/Dsg3, Dsc1/Dsc1) or heterophilic combinations (Dsg1/Dsc1 and Dsg3/Dsc1). (B) Inhibition experiments showing heterophilic aggregation of green Dsc1-coated beads with red Dsg1 (top panels) or Dsg3- coated beads (bottom panels) pre-incubated with purified IgG4 fractions from FS patients or from a healthy donor (see methods for details). (C) Dose-dependence of IgG4 Fab mediated inhibition of Dsg1/Dsc1 bead aggregation from donors FS 1 and FS17. Scale bar 0.5mm.

    Article Snippet: Commercially available human IgG4 (Sigma Aldrich, St Louis, MO, USA) was injected as a control.

    Techniques: Inhibition, Incubation, Purification

    FS IgG4 autoantibodies mainly recognize Dsg1 EC1 domain ( A ) The EC1 domain of Dsg1 (blue) was grafted on three molecular backbones: Dsg3 (red domains), Dsg4 (white domains) and Dsc1 (green domains). We also grafted the EC1 domain of Dsg4 on a Dsg1 backbone (blue domains) as a control hybrid. These constructs were expressed in the baculovirus system and the recombinant proteins were used to test against FS sera. ( B ) Affinity-purified FS IgG4 on the llama anti-human IgG4 column was tested against chimeric Dsg1-EC1/Dsg3 bb in the presence of calcium and EDTA. A stock of IgG4 (0.5 mg/ml) of each patient at a dilution of 1:300 was tested on microtiter wells coated with 50 ng/well of Dsg1-EC1/Dsg3 bb. Peroxidase-labeled mouse anti-human IgG4 (Southern Biotech) at a dilution of 1:2,000 was used as an indicator. The results are expressed in index values. Similar results were observed when testing the FS IgG4 on plates coated with Dsg1-EC1/Dsg4 bb (data not shown). ( C) Affinity-purified FS IgG4 on the llama anti-human IgG4 column was tested against chimeric Dsg1-EC1/Dsg4 bb (blue), Dsg4-EC1/Dsg1 bb (red), wDsg4 (green) and wDsc1 (purple) in the presence of calcium. A stock of IgG4 (0.5 mg/ml) of each patient at a dilution of 1:300 was tested on microtiter wells coated with 50 ng/well of the chimeric protein. Peroxidase-labeled mouse anti-human IgG4 (Southern Biotech) at the dilution of 1:2,000 was used as an indicator. The results are expressed in index values (Scale of -20 to 100 is shown in the figure).

    Journal: Journal of autoimmunity

    Article Title: Pathogenic IgG4 autoantibodies from endemic pemphigus foliaceus recognize a desmoglein-1 conformational epitope

    doi: 10.1016/j.jaut.2017.12.017

    Figure Lengend Snippet: FS IgG4 autoantibodies mainly recognize Dsg1 EC1 domain ( A ) The EC1 domain of Dsg1 (blue) was grafted on three molecular backbones: Dsg3 (red domains), Dsg4 (white domains) and Dsc1 (green domains). We also grafted the EC1 domain of Dsg4 on a Dsg1 backbone (blue domains) as a control hybrid. These constructs were expressed in the baculovirus system and the recombinant proteins were used to test against FS sera. ( B ) Affinity-purified FS IgG4 on the llama anti-human IgG4 column was tested against chimeric Dsg1-EC1/Dsg3 bb in the presence of calcium and EDTA. A stock of IgG4 (0.5 mg/ml) of each patient at a dilution of 1:300 was tested on microtiter wells coated with 50 ng/well of Dsg1-EC1/Dsg3 bb. Peroxidase-labeled mouse anti-human IgG4 (Southern Biotech) at a dilution of 1:2,000 was used as an indicator. The results are expressed in index values. Similar results were observed when testing the FS IgG4 on plates coated with Dsg1-EC1/Dsg4 bb (data not shown). ( C) Affinity-purified FS IgG4 on the llama anti-human IgG4 column was tested against chimeric Dsg1-EC1/Dsg4 bb (blue), Dsg4-EC1/Dsg1 bb (red), wDsg4 (green) and wDsc1 (purple) in the presence of calcium. A stock of IgG4 (0.5 mg/ml) of each patient at a dilution of 1:300 was tested on microtiter wells coated with 50 ng/well of the chimeric protein. Peroxidase-labeled mouse anti-human IgG4 (Southern Biotech) at the dilution of 1:2,000 was used as an indicator. The results are expressed in index values (Scale of -20 to 100 is shown in the figure).

    Article Snippet: Commercially available human IgG4 (Sigma Aldrich, St Louis, MO, USA) was injected as a control.

    Techniques: Construct, Recombinant, Affinity Purification, Labeling

    Normal donors from endemic, non-endemic and US do not recognize Dsg1-EC1/Dsg3 bb The sera of three sets of normal individuals were tested by ELISA against Dsg1-EC1/Dsg3 bb coated plates. The sera were tested at 1:300 dilution and mouse anti-human IgG4 was used at the 1:2000 dilution. The plates were coated with 100 ng/well of Dsg1-Ec1/Dsg3 bb. The results are expressed in index values (scale of -20 to 100 is shown in the figure). Panel A ] compared with the reaction of FS-1 included as a positive control. Normal human serum (NHS) from the US served as a negative control. Panel B shows the lack of reactivity of the sera of twenty nine normal individuals (C1 to C-29) from non-endemic areas of Brazil (urban inhabitants of the cities of Sao Paulo and Belho Horizonte) compared with FS-1 and NHS. Donor C-17 was found later to be a case of FS undergoing therapy and in clinical remission. Panel C shows the lack of reactivity of the sera of thirty normal individuals (C-1 to C-30) from the UNC Blood Bank compared with FS-1 and NHS.

    Journal: Journal of autoimmunity

    Article Title: Pathogenic IgG4 autoantibodies from endemic pemphigus foliaceus recognize a desmoglein-1 conformational epitope

    doi: 10.1016/j.jaut.2017.12.017

    Figure Lengend Snippet: Normal donors from endemic, non-endemic and US do not recognize Dsg1-EC1/Dsg3 bb The sera of three sets of normal individuals were tested by ELISA against Dsg1-EC1/Dsg3 bb coated plates. The sera were tested at 1:300 dilution and mouse anti-human IgG4 was used at the 1:2000 dilution. The plates were coated with 100 ng/well of Dsg1-Ec1/Dsg3 bb. The results are expressed in index values (scale of -20 to 100 is shown in the figure). Panel A ] compared with the reaction of FS-1 included as a positive control. Normal human serum (NHS) from the US served as a negative control. Panel B shows the lack of reactivity of the sera of twenty nine normal individuals (C1 to C-29) from non-endemic areas of Brazil (urban inhabitants of the cities of Sao Paulo and Belho Horizonte) compared with FS-1 and NHS. Donor C-17 was found later to be a case of FS undergoing therapy and in clinical remission. Panel C shows the lack of reactivity of the sera of thirty normal individuals (C-1 to C-30) from the UNC Blood Bank compared with FS-1 and NHS.

    Article Snippet: Commercially available human IgG4 (Sigma Aldrich, St Louis, MO, USA) was injected as a control.

    Techniques: Enzyme-linked Immunosorbent Assay, Positive Control, Negative Control

    Epitopes recognized by FS IgG4 autoantibodies Five Dsg1 peptides were bound by FS IgG4 from 20 patients: Propeptide (W41 to R45) (45%); EC1 (A 129 to R 144 ) (95%); EC2 (Q 201 to R 213 ) (85%); EC4 (Y 460 to R 473 ) (40%) and EC5 (D 538 to F 546 ) (25%). This investigation focused on the EC1 epitope.

    Journal: Journal of autoimmunity

    Article Title: Pathogenic IgG4 autoantibodies from endemic pemphigus foliaceus recognize a desmoglein-1 conformational epitope

    doi: 10.1016/j.jaut.2017.12.017

    Figure Lengend Snippet: Epitopes recognized by FS IgG4 autoantibodies Five Dsg1 peptides were bound by FS IgG4 from 20 patients: Propeptide (W41 to R45) (45%); EC1 (A 129 to R 144 ) (95%); EC2 (Q 201 to R 213 ) (85%); EC4 (Y 460 to R 473 ) (40%) and EC5 (D 538 to F 546 ) (25%). This investigation focused on the EC1 epitope.

    Article Snippet: Commercially available human IgG4 (Sigma Aldrich, St Louis, MO, USA) was injected as a control.

    Techniques:

    The 16-residue alignment among different desmosomal proteins The 16-residue peptide bound by 19/20 FS IgG4 (A 129 to R 144 ) on Dsg1 was aligned with similar peptides from Dsg2, Dsg3, Dsg4, Dsc1 and E-Cadherin. The Dsg4 peptide shows two residues, R 133 and E 135 that are different from the Dsg1 counterpart. Human Dsg4 does not bind FS IgG or IgG4 autoantibodies.

    Journal: Journal of autoimmunity

    Article Title: Pathogenic IgG4 autoantibodies from endemic pemphigus foliaceus recognize a desmoglein-1 conformational epitope

    doi: 10.1016/j.jaut.2017.12.017

    Figure Lengend Snippet: The 16-residue alignment among different desmosomal proteins The 16-residue peptide bound by 19/20 FS IgG4 (A 129 to R 144 ) on Dsg1 was aligned with similar peptides from Dsg2, Dsg3, Dsg4, Dsc1 and E-Cadherin. The Dsg4 peptide shows two residues, R 133 and E 135 that are different from the Dsg1 counterpart. Human Dsg4 does not bind FS IgG or IgG4 autoantibodies.

    Article Snippet: Commercially available human IgG4 (Sigma Aldrich, St Louis, MO, USA) was injected as a control.

    Techniques:

    FS IgG4 autoantibodies are pathogenic The IgG4 fraction from FS1 serum (from patient #1 in the set of 20 tested, FS1 is used throughout the paper) was purified on a llama anti-human IgG4 Sepharose affinity column. The bound fraction contained 97% pure IgG4. The unbound fraction contained non-IgG4 subclasses with IgG1 representing the majority (92%). The IgG4 and non-IgG4 fractions were dialyzed, concentrated and injected subcutaneously into neonatal mice. A. Mice injected with FS1 IgG4 (left) developed extensive blistering, shown as fine wrinkling of the epidermis (induced by slight friction or pinching, the so-called Nikolsky’s sign). B. On histological examination large sheets of epidermis were separated from the naked dermis (x200) and direct immunofluorescence shows bound IgG4 to the roof of the split (x100). Mice receiving a higher dose of non-IgG4 fraction showed no disease (right).

    Journal: Journal of autoimmunity

    Article Title: Pathogenic IgG4 autoantibodies from endemic pemphigus foliaceus recognize a desmoglein-1 conformational epitope

    doi: 10.1016/j.jaut.2017.12.017

    Figure Lengend Snippet: FS IgG4 autoantibodies are pathogenic The IgG4 fraction from FS1 serum (from patient #1 in the set of 20 tested, FS1 is used throughout the paper) was purified on a llama anti-human IgG4 Sepharose affinity column. The bound fraction contained 97% pure IgG4. The unbound fraction contained non-IgG4 subclasses with IgG1 representing the majority (92%). The IgG4 and non-IgG4 fractions were dialyzed, concentrated and injected subcutaneously into neonatal mice. A. Mice injected with FS1 IgG4 (left) developed extensive blistering, shown as fine wrinkling of the epidermis (induced by slight friction or pinching, the so-called Nikolsky’s sign). B. On histological examination large sheets of epidermis were separated from the naked dermis (x200) and direct immunofluorescence shows bound IgG4 to the roof of the split (x100). Mice receiving a higher dose of non-IgG4 fraction showed no disease (right).

    Article Snippet: Commercially available human IgG4 (Sigma Aldrich, St Louis, MO, USA) was injected as a control.

    Techniques: Purification, Affinity Column, Injection, Mouse Assay, Immunofluorescence

    Onchocerca volvulus adult worm (OvAg) and recombinant Ov16 antigen-specific IgG4 reactivity (optical densities; OD) in participants and positive and negative IgG4 responses in age groups. In A and C the data on antigen-specific IgG4 reactivity are shown as mean optical densities (ODs) with 95% confidence intervals for the means (diamonds). The data presented in box plots show the median OD per age group with the 25% and 75% quartiles and the 1.5x of the interquartile range. In B and D the antigen-specific-IgG4 positive and negative responses in age groups are indicated (in %).

    Journal: PLoS Neglected Tropical Diseases

    Article Title: Onchocerca volvulus infection and serological prevalence, ocular onchocerciasis and parasite transmission in northern and central Togo after decades of Simulium damnosum s.l. vector control and mass drug administration of ivermectin

    doi: 10.1371/journal.pntd.0006312

    Figure Lengend Snippet: Onchocerca volvulus adult worm (OvAg) and recombinant Ov16 antigen-specific IgG4 reactivity (optical densities; OD) in participants and positive and negative IgG4 responses in age groups. In A and C the data on antigen-specific IgG4 reactivity are shown as mean optical densities (ODs) with 95% confidence intervals for the means (diamonds). The data presented in box plots show the median OD per age group with the 25% and 75% quartiles and the 1.5x of the interquartile range. In B and D the antigen-specific-IgG4 positive and negative responses in age groups are indicated (in %).

    Article Snippet: After using PBS-Tween20 (Sigma P3563) for washing, an anti-human IgG4 horseradish peroxidase conjugated monoclonal antibody (Thermo Fisher Scientific, no. A10654) was added (dilution 1:500) for 1.5 hours.

    Techniques: Recombinant

    Evaluating IgG subclass response to the 39041 epitope ( n = 6). The Kruskal-Wallis test was used for analyzing differences between each IgG subclass response in P. vivax -exposed individuals' samples. ** p

    Journal: Frontiers in Cellular and Infection Microbiology

    Article Title: The in Vitro Antigenicity of Plasmodium vivax Rhoptry Neck Protein 2 (PvRON2) B- and T-Epitopes Selected by HLA-DRB1 Binding Profile

    doi: 10.3389/fcimb.2018.00156

    Figure Lengend Snippet: Evaluating IgG subclass response to the 39041 epitope ( n = 6). The Kruskal-Wallis test was used for analyzing differences between each IgG subclass response in P. vivax -exposed individuals' samples. ** p

    Article Snippet: A 1:1,000 dilution of monoclonal anti-human IgG1–biotin antibody produced in mouse (clone 8c/6-39), 1:15,000 monoclonal anti-human IgG2–biotin antibody produced in mouse (clone HP-6014), 1:40,000 monoclonal anti-human IgG3–biotin antibody produced in mouse (clone HP-6050) and 1:60,000 anti-human IgG4–biotin antibody, mouse monoclonal (clone HP-6025) (Sigma Aldrich) were used.

    Techniques:

    IgG antibody response to Pv RON2 B epitopes by endemic area. Significant differences (calculated by Mann-Whitney test) between samples from Colombia's Chocó ( n = 13) and Córdoba ( n = 17) departments are shown. The dashed line indicates the cut-off point for seropositive samples. * p

    Journal: Frontiers in Cellular and Infection Microbiology

    Article Title: The in Vitro Antigenicity of Plasmodium vivax Rhoptry Neck Protein 2 (PvRON2) B- and T-Epitopes Selected by HLA-DRB1 Binding Profile

    doi: 10.3389/fcimb.2018.00156

    Figure Lengend Snippet: IgG antibody response to Pv RON2 B epitopes by endemic area. Significant differences (calculated by Mann-Whitney test) between samples from Colombia's Chocó ( n = 13) and Córdoba ( n = 17) departments are shown. The dashed line indicates the cut-off point for seropositive samples. * p

    Article Snippet: A 1:1,000 dilution of monoclonal anti-human IgG1–biotin antibody produced in mouse (clone 8c/6-39), 1:15,000 monoclonal anti-human IgG2–biotin antibody produced in mouse (clone HP-6014), 1:40,000 monoclonal anti-human IgG3–biotin antibody produced in mouse (clone HP-6050) and 1:60,000 anti-human IgG4–biotin antibody, mouse monoclonal (clone HP-6025) (Sigma Aldrich) were used.

    Techniques: MANN-WHITNEY

    IgG antibody response against four Pv RON2 B-cell epitopes ( n = 30). Seropositive samples were those above the cut-off point (0.218, dotted line), calculated as control group's mean plus two standard deviations. The Kruskal-Wallis test was used for analyzing differences between each B-epitopes response in P. vivax -exposed individuals' samples. * p

    Journal: Frontiers in Cellular and Infection Microbiology

    Article Title: The in Vitro Antigenicity of Plasmodium vivax Rhoptry Neck Protein 2 (PvRON2) B- and T-Epitopes Selected by HLA-DRB1 Binding Profile

    doi: 10.3389/fcimb.2018.00156

    Figure Lengend Snippet: IgG antibody response against four Pv RON2 B-cell epitopes ( n = 30). Seropositive samples were those above the cut-off point (0.218, dotted line), calculated as control group's mean plus two standard deviations. The Kruskal-Wallis test was used for analyzing differences between each B-epitopes response in P. vivax -exposed individuals' samples. * p

    Article Snippet: A 1:1,000 dilution of monoclonal anti-human IgG1–biotin antibody produced in mouse (clone 8c/6-39), 1:15,000 monoclonal anti-human IgG2–biotin antibody produced in mouse (clone HP-6014), 1:40,000 monoclonal anti-human IgG3–biotin antibody produced in mouse (clone HP-6050) and 1:60,000 anti-human IgG4–biotin antibody, mouse monoclonal (clone HP-6025) (Sigma Aldrich) were used.

    Techniques: