goat anti rabbit  (Bio-Rad)

Journal: Journal of Bacteriology

Article Title: Identification of the ?4514 Regulatory Region, a Developmental Promoter of Myxococcus xanthus That Is Transcribed In Vitro by the Major Vegetative RNA Polymerase

doi: 10.1128/JB.184.12.3348-3359.2002

Figure Lengend Snippet: Western blot analysis of σ A in growing and developing M. xanthus cells. DK1622 cells were collected at 0, 9, 12, 15, 18, and 24 h after the onset of development (lanes 1 to 6), resuspended in sample buffer, and boiled for 5 min. Equal amounts of M. xanthus protein, as judged by measuring total protein in sonic lysates of the same samples, were separated by SDS-10% PAGE and electrotransferred to a nitrocellulose filter, which was incubated with a polyclonal antiserum to B. subtilis σ 43 and visualized with goat anti-rabbit immunoglobulin G-horseradish peroxidase. The σ A is indicated by an arrow, and protein standards are marked in kilodaltons.

Article Snippet: Immunodetection with goat anti-rabbit immunoglobulin G-horseradish peroxidase (Bio-Rad) and chemiluminescence (ECL; Amersham Pharmacia Biotech) was performed according to the manufacturer's instructions.

Techniques: Western Blot, Polyacrylamide Gel Electrophoresis, Incubation

Journal: The Journal of Cell Biology

Article Title: Role of P-Selectin Cytoplasmic Domain in Granular Targeting In Vivo and in Early Inflammatory Responses

doi:

Figure Lengend Snippet: Localization of ΔCT–P-selectin in platelets by electron microscopy. Indirect immunogold labeling of P-selectin was performed in resting platelets from wild-type ( WT ) and ΔCT mice. Ultrathin frozen sections were stained with a rabbit antibody against P-selectin and visualized with a goat anti–rabbit IgG conjugated to 10-nm colloidal gold particles. The majority of the gold particles are associated with α-granules ( arrowheads ) in both wild-type and ΔCT platelets. A small amount of labeling was seen on the plasma membrane in both genotypes. Bars, 200 nm.

Article Snippet: The antibody bound to P-selectin was detected with a horseradish peroxidase-conjugated goat anti–rabbit IgG (Bio-Rad, Hercules, CA) and an enhanced chemiluminescence kit ( Sigma Chemical Co. , St. Louis, MO).

Techniques: Electron Microscopy, Labeling, Mouse Assay, Staining

Journal: The Journal of Cell Biology

Article Title: Role of P-Selectin Cytoplasmic Domain in Granular Targeting In Vivo and in Early Inflammatory Responses

doi:

Figure Lengend Snippet: Flow cytometry analysis of P-selectin expression on platelets. Wild-type and ΔCT platelets were stained for membrane P-selectin and analyzed by flow cytometry. In the resting state, platelets of both genotypes displayed virtually no P-selectin on their plasma membranes. Thrombin activation induced in wild-type as well as in mutant platelets a similar increase in mean fluorescence with ∼90% of P-selectin–positive platelets. Representative histograms are shown. Shaded area , negative control staining with only the FITC-conjugated goat anti–rabbit IgG.

Article Snippet: The antibody bound to P-selectin was detected with a horseradish peroxidase-conjugated goat anti–rabbit IgG (Bio-Rad, Hercules, CA) and an enhanced chemiluminescence kit ( Sigma Chemical Co. , St. Louis, MO).

Techniques: Flow Cytometry, Cytometry, Expressing, Staining, Activation Assay, Mutagenesis, Fluorescence, Negative Control

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Tissue Inhibitor of Metalloproteinase-1 (TIMP-1) Promotes PMN Pericellular Proteolysis by Anchoring Matrix Metalloproteinase-8 and -9 to PMN Surfaces

doi: 10.4049/jimmunol.1801466

Figure Lengend Snippet: ProMMP-8 and proMMP-9 share binding sites on the surface of PMNs: In A and B , PMNs from Mmp-8 −/− x Mmp-9 −/− mice were activated for 15 min at 37°C with 10 −6 M PAF followed by 10 −6 M fMLP to induce surface expression of Timp-1. In A , the activated PMNs were then pre-incubated for 45 min at 4°C with or without exogenous full length murine proMmp-8 (50–400 nM). Cells were then incubated for an additional 60 min at 4°C with 100 nM exogenous murine proMMP-9. Cells were then washed and immunostained with rabbit anti-murine Mmp-9 IgG, or non-immune rabbit IgG followed by goat anti-rabbit F(ab) 2 -conjugated to Alexa 488 to detect bound pro and active Mmp-9. In B , the PAF- and fMLP- activated and PMNs were pre-incubated for 45 min at 4°C with or without exogenous full length murine proMmp-9 (50–400 nM). Cells were then incubated for an additional 60 min at 4°C with 100 nM exogenous full length murine proMMP-8. Cells were then immunostained with rabbit anti-murine Mmp-8 IgG, or non-immune rabbit IgG followed by goat anti-rabbit F(ab) 2 -conjugated to Alexa 488 to detect bound pro and active Mmp-8. In A and B , images of immunostained cells were captured and surface-bound pro and active Mmp-9 (in A ) or pro and active Mmp-8 (in B ) were quantified using MetaMorph software. Data are mean + SEM (n = 150–200 cells/group). Data were analyzed using a One-Way ANOVA followed by pair-wise testing with two-tailed Student’s t-tests. Asterisk indicates P

Article Snippet: Briefly, samples were subjected to electrophoresis on 8% SDS-PAGE under reducing conditions, proteins transferred to PVDF membranes, and probed with polyclonal rabbit anti-murine Mmp-9 IgG (AB19047) or rabbit anti-murine Mmp-8 IgG (Ab19045), followed by goat anti-rabbit IgG conjugated to horse radish peroxidase (Biorad, Hercules, CA), and a chemiluminescence detection system following the manufacturer’s instructions.

Techniques: Binding Assay, Mouse Assay, Expressing, Incubation, Software, Two Tailed Test

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Tissue Inhibitor of Metalloproteinase-1 (TIMP-1) Promotes PMN Pericellular Proteolysis by Anchoring Matrix Metalloproteinase-8 and -9 to PMN Surfaces

doi: 10.4049/jimmunol.1801466

Figure Lengend Snippet: TIMP-1 is co-localized with MMP-8 and MMP-9 on the surface of activated human PMNs: Human PMNs were activated at 37°C with PAF at 10 −7 M for 15 min followed by fMLP at 10 −7 M for 30 min. Cells were double immunostained with Alexa 546 and murine anti-MMP-9 IgG ( A , second panel), or murine anti-human MMP-8 IgG ( B , second panel) or non-immune murine (Ms) IgG ( C , second panel) and with Alexa 488 and rabbit anti-TIMP-1 IgG ( A and B third panels) or non-immune rabbit (Rb) IgG ( C , third panel). The anti-MMP-8 and anti-MMP-9 IgGs used recognize both pro and active forms of these MMPs. Cells were examined using a Normarski objective ( A-C , left panels) and co-localization of MMPs and TIMP-1 on the surface of the activated PMNs was assessed by confocal microscopy (see merged images in the right panels for A-C ). The white bars are 5 microns in length. Note the strong co-localization of TIMP-1 with both MMP-8 and MMP-9 on the surface of activated PMNs. The results shown are representative of at least 3 different PMN preparations.

Article Snippet: Briefly, samples were subjected to electrophoresis on 8% SDS-PAGE under reducing conditions, proteins transferred to PVDF membranes, and probed with polyclonal rabbit anti-murine Mmp-9 IgG (AB19047) or rabbit anti-murine Mmp-8 IgG (Ab19045), followed by goat anti-rabbit IgG conjugated to horse radish peroxidase (Biorad, Hercules, CA), and a chemiluminescence detection system following the manufacturer’s instructions.

Techniques: Confocal Microscopy

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Tissue Inhibitor of Metalloproteinase-1 (TIMP-1) Promotes PMN Pericellular Proteolysis by Anchoring Matrix Metalloproteinase-8 and -9 to PMN Surfaces

doi: 10.4049/jimmunol.1801466

Figure Lengend Snippet: Exogenous Timp-1 reconstitutes the binding of exogenous proMmp-9 and proMmp-8 to Timp-1 −/− PMNs: In A , PAF- and fMLP-activated PMNs from Mmp-9 −/− and Timp-1 −/− mice were incubated with or without 50–400 nM exogenous murine proMmp-9, and then immunostained with rabbit anti-Mmp-9 IgG, or non-immune rabbit IgG followed by goat anti-rabbit F(ab) 2 -conjugated to Alexa 488. In B , PAF- and fMLP-activated PMNs from Mmp-8 −/− and Timp-1 −/− mice were incubated with or without 50–400 nM exogenous murine proMmp-8, and then immunostained with rabbit anti-Mmp-8 IgG, or non-immune rabbit IgG followed by goat- anti-rabbit F(ab) 2 -conjugated to Alexa 488. In A-B , the antibodies used detect both the pro and active forms of the Mmps. Data are mean + SEM; n = 150–200 cells per group. Data were analyzed using a One-Way ANOVA followed by pair-wise testing with two-tailed Student’s t-tests. Asterisk indicates P

Article Snippet: Briefly, samples were subjected to electrophoresis on 8% SDS-PAGE under reducing conditions, proteins transferred to PVDF membranes, and probed with polyclonal rabbit anti-murine Mmp-9 IgG (AB19047) or rabbit anti-murine Mmp-8 IgG (Ab19045), followed by goat anti-rabbit IgG conjugated to horse radish peroxidase (Biorad, Hercules, CA), and a chemiluminescence detection system following the manufacturer’s instructions.

Techniques: Binding Assay, Mouse Assay, Incubation, Two Tailed Test

Journal: Vaccine

Article Title: Recombinant Attenuated Listeria monocytogenes Vaccine Expressing Francisella tularensis IglC Induces Protection in Mice Against Aerosolized Type A F. tularensis

doi: 10.1016/j.vaccine.2008.12.014

Figure Lengend Snippet: Antibody response induced by intradermal immunization with recombinant L. monocytogenes. Groups of 5 mice were sham-immunized or immunized i.d. twice at weeks 0 and 4 with LVS, vector control (rLmΔactA), or recombinant L. monocytog enes vaccines expressing KatG (rLm/katG), IglC (rLm/iglC), or the combination of both vaccines. At week 8, mice were euthanized and sera were collected to measure the antibody (IgG) response by ELISA. For measuring antibody response to KatG (A), sera were diluted 1:50. For measuring antibody response to IglC (B), sera were diluted 1:25. The antibody responses are presented as the Absorbance at 414 nm (A 414nm ) in the presence of antigen divided by the A 414nm in the absence of antigen for each individual mouse. Bars represent the mean of the antibody titer from the 5 mice per group. Asterisks indicate that the difference between the antibody response of mice in the indicated vaccinated group and mice in the sham-vaccinated group was significant. *, P

Article Snippet: Protein expression by recombinant L. monocytogenes was analyzed by Western blotting using rabbit polyclonal antibodies specific to AcpA, Bfr, DnaK, GroEL, IglC, KatG, or Pld at a dilution of 1:5,000 and subsequently horseradish peroxidase-conjugated goat anti-rabbit immunoglobulin G (Bio-Rad) at a dilution of 1:25,000.

Techniques: Recombinant, Mouse Assay, Plasmid Preparation, Expressing, Enzyme-linked Immunosorbent Assay