Journal: Journal of Clinical Medicine

Article Title: Effects of Caloric Restriction with Protein Supplementation on Plasma Protein Profiles in Middle-Aged Women with Metabolic Syndrome—A Preliminary Open Study

doi: 10.3390/jcm8020195

Figure Lengend Snippet: Heat maps showing different baseline and postintervention protein plasma levels. A significant correlation was observed with the selected increment clinical variables between baseline and postintervention in the CR ( A ) and CRPS ( B ) groups. The left red–blue scale heatmaps show the estimates of Spearman’s correlation coefficients, whereas the right yellow-scale heat maps show the p -values for the same protein–clinical variable pairs. AG: android/gynoid fat ratio, ALB: albumin, TBF: total body fat, BMI: body mass index, CRP: C-reactive protein, FBG: fasting blood glucose, HOMA: homeostatic model assessment for insulin resistance, IL6: interleukin 6, LDL-C: low-density lipoprotein cholesterol, MONO: mononuclear, NEU: neutrophil, PLA: platelet, QUICKI: quantitative insulin sensitivity check index, TC: total cholesterol, WBC: white blood cell, A2M: alpha-2-macroglobulin, C1RL: Complement C1r subcomponent–like protein, C4BPA: C4b-binding protein alpha chain, C6: complement component C6, C8G: complement component C8 gamma chain, Protein S: vitamin K-dependent protein S, CR: caloric restriction, CRPS: caloric restriction with protein supplementation.

Article Snippet: Commercial available plasma C4b-binding protein (C4BP), complement component C6 (C6), complement component C8 gamma chain (C8G), and vitamin K–dependent protein S (PROS) were respectively measured using the following commercial ELISA kits: (1) C4BP ELISA kit (#EC2202-1, Assaypro, St. Charles, MO, USA); (2) C6 ELISA kit (#EC6101-1, Assaypro, St. Charles, MO, USA); (3) C8G ELISA kit (#EC8120-1, Assaypro, St. Charles, MO, USA); and (4) PROS ELISA kit (#AB190808, Abcam, Cambridge, UK).

Techniques: Clinical Proteomics, Binding Assay

Journal: Molecular Neurobiology

Article Title: Novel Exosome Biomarker Candidates for Alzheimer’s Disease Unravelled Through Mass Spectrometry Analysis

doi: 10.1007/s12035-022-02762-1

Figure Lengend Snippet: AACT exosomal levels in dementia and AD cases monitored by distinct antibody-based approaches. AACT levels were assessed through immunoblot analysis or commercial ELISA assays in serum-derived exosomes from Controls (CDR = 0 and MMSE −) and individuals with dementia (CDR ≥ 1 and MMSE +) from UA-cohort ( a , d ), and AD clinically diagnosed cases from UA-cohort ( b , e ) or UMG-cohort ( c , f ). For WB, each point represents the relative densitometry ratio. For ELISA, each point represents the mean concentration value obtained for each individual. The solid horizontal line shows mean, and error bars indicates standard deviations. Abbreviations: AD, Alzheimer’s disease; C, Controls; CDR, Clinical Dementia Rate; MMSE, Mini-Mental State Examination. * p ≤ 0.05

Article Snippet: Further, AACT and C4BPα levels were also evaluated by enzyme-linked immunosorbent assay (ELISA), in serum-derived exosomes of the same individuals, using the commercial Human AACT ELISA Kit (ab217779; Abcam) or the Human C4 binding protein A ELISA Kit (NBP2-60,550; Novus Biologicals), according to manufacturer’s instructions.

Techniques: Western Blot, Enzyme-linked Immunosorbent Assay, Derivative Assay, Concentration Assay

Journal: Frontiers in Oncology

Article Title: Establishment, characterization, and drug screening of low-passage patient individual non-small cell lung cancer in vitro models including the rare pleomorphic subentity

doi: 10.3389/fonc.2023.1089681

Figure Lengend Snippet: Antibodies used for flow cytometry.

Article Snippet: LYPD3 , APC , Sino Biological Europe GmbH, Düsseldorf, Germany , 11836-H08H.

Techniques: Cytometry

Journal: Frontiers in Oncology

Article Title: Establishment, characterization, and drug screening of low-passage patient individual non-small cell lung cancer in vitro models including the rare pleomorphic subentity

doi: 10.3389/fonc.2023.1089681

Figure Lengend Snippet: Flow cytometry. Histogram overlays of unstained controls (dotted lines for all three cell lines) and measurements for HROLu55 (green), HROLu22 (blue), and HROBML01 (red) for the epitopes CD326, PD-L1, EGFR, CD26, LYPD3, DSG3, CCD59, CD27, and CD90 are shown.

Article Snippet: LYPD3 , APC , Sino Biological Europe GmbH, Düsseldorf, Germany , 11836-H08H.

Techniques: Flow Cytometry

Journal: Frontiers in Immunology

Article Title: Toxoplasma gondii Recruits Factor H and C4b-Binding Protein to Mediate Resistance to Serum Killing and Promote Parasite Persistence in vivo

doi: 10.3389/fimmu.2019.03105

Figure Lengend Snippet: Toxoplasma gondii recruits AP regulator Factor H and CP/LP regulator C4b-binding protein to the parasite surface. 1 × 10 6 Type II ME49 parasites were incubated in 10% NHS for 0–60 min at 37°C. Western blots (left panels) of C4BP (A) (rabbit α-human C4BP, AssayPro 1:500) and FH (B) (goat α-human Factor H, CompTech 1:20,000) binding. Serum or purified protein was used as a positive control and heat inactivated serum (hiNHS) was used as a negative control. Blots were stripped and re-probed with anti-SRS29B (SAG1) for loading control. Images are from one representative of three independent experiments with similar results. Right panels (A,B) represent flow cytometric assays of C4BP (A) and FH (B) binding to the parasite surface for 0–60 min. Heat inactivated NHS (hiNHS) serum was used as a negative control. Flow cytometry data are shown as mean ± SEM from three independently performed experiments.

Article Snippet: The following antibody dilutions were used: rabbit anit-SRS29B (formerly SAG1, 30 kDa) 1:5,000, rabbit anti-C4BP (alpha chain, 70 kDa) (AssayPro) 1:1,000, goat anti-C3 (C3, C3b, iC3b) and goat anti-Factor H (full length protein, 155 kDa) antibodies (CompTech) were both used at 1:20,0000, anti-rabbit HRP 1:10,000 (Sigma), and anti-goat HRP 1:5,000 (Santa Cruz Biotechnology, Inc.).

Techniques: Binding Assay, Incubation, Western Blot, Purification, Positive Control, Negative Control, Flow Cytometry

Journal: Frontiers in Immunology

Article Title: Toxoplasma gondii Recruits Factor H and C4b-Binding Protein to Mediate Resistance to Serum Killing and Promote Parasite Persistence in vivo

doi: 10.3389/fimmu.2019.03105

Figure Lengend Snippet: Factor H and C4b-binding protein contribute to serum resistance. Factor H (FH) and C4b-binding protein (C4BP) were blocked by pre-incubating 10% NHS with 1:100 or 1:400 dilution of goat α-human FH (CompTech) or rabbit α-human C4BP (AssayPro) for 1 h on ice before adding to 1 × 10 6 parasites and incubating for 60 min at 37°C. Flow cytometric analysis of C5b-9 formation (A) and parasite viability (B) after 60' in 10% NHS blocked with 1:100 or 1:400 of α-C4BP (gray bars) α-FH (open bars) antibodies. 10% heat inactivated serum (hiNHS) was used a negative control. Flow cytometry data are shown as mean ± SEM from three independently performed experiments. Significant differences between the compared groups was determined using multiple Student's t test with Holm-Sidak correction for multiple comparisons, * p < 0.05, *** p < 0.001, **** p < 0.0001.

Article Snippet: The following antibody dilutions were used: rabbit anit-SRS29B (formerly SAG1, 30 kDa) 1:5,000, rabbit anti-C4BP (alpha chain, 70 kDa) (AssayPro) 1:1,000, goat anti-C3 (C3, C3b, iC3b) and goat anti-Factor H (full length protein, 155 kDa) antibodies (CompTech) were both used at 1:20,0000, anti-rabbit HRP 1:10,000 (Sigma), and anti-goat HRP 1:5,000 (Santa Cruz Biotechnology, Inc.).

Techniques: Binding Assay, Negative Control, Flow Cytometry

Journal: PLoS ONE

Article Title: Ficolin-2 Defends against Virulent Mycobacteria Tuberculosis Infection In Vivo , and Its Insufficiency Is Associated with Infection in Humans

doi: 10.1371/journal.pone.0073859

Figure Lengend Snippet: (A) Ficolin-2/ficolin A inhibited Mtb H37Rv infection of human lung A549 cells in vitro . pcDNA3.1-ficolin-2 or pVAX-1-ficolin-A plasmids were transfected into A549 cells for 48 h. Rhodamine B-labeled Mtb H37Rv (RhoB-H37Rv) was mixed with ficolin-2- or ficolin A-transfected human lung A549 cells with or without ManLAM, and the cells were analyzed using flow cytometry. (B) Ficolin-2 stimulated opsonization of macrophages. RhoB-H37Rv bacteria were incubated with 20 µg of GST, GST-ficolin-2, GST-ficolin-2 plus CytB, or CytB, respectively, at 4°C for 30 min. The uptake of the H37Rv strain (detected as Rho-H37Rv) by macrophages (detected as FITC-F4/80) was analyzed using flow cytometry. The results shown are representative of three independent experiments. (C) The binding of ficolin-2 with the H37Rv strain leads to the activation of the lectin complement pathway. GST-ficolin-2 or GST proteins were incubated with Mtb H37Rv in the presence of human serum and C4 protein, and C4c deposits on the bacteria were measured as described in the Materials and Methods. The data shown are the mean ± SEM of three independent experiments GST-ficolin-2 group vs. GST group and control group, * p < 0.05. The data were analyzed by ANOVA method.

Article Snippet: The bacteria were then washed thoroughly, and 0.1 μg of purified human C4 protein (Diagnostic Biosystems) was added and incubated at 37°C for 1.5 h. The bacteria were washed again, and FITC-conjugated rabbit anti-human C4c (Diagnostic Biosystems) (1:100 dilution) was added and incubated at room temperature for 30 min.

Techniques: Infection, In Vitro, Transfection, Labeling, Flow Cytometry, Bacteria, Incubation, Binding Assay, Activation Assay, Control