Journal: Immunology letters

Article Title: Endogenous Retrovirus Gag Antigen and Its Gene Variants Are Unique Autoantigens Expressed in the Pancreatic Islets of Non-obese Diabetic Mice

doi: 10.1016/j.imlet.2020.04.007

Figure Lengend Snippet: Detection of Gag antigen in pancreatic islets. A and B: Frozen sections of pancreas from female NOD (i&iv), B6(ii&v) and (NODxB6)F1 (iii&vi) mice (8-12 week-old) were stained with R187 anti-Gag mAb, followed by HRP-labeled (A) or AP-labeled (B) anti-rat IgG secondary antibodies and color development using AEC (A) or BCIP/NBT (B) substrate, respectively. C: RT-PCR was performed using total RNA (genomic DNA-free) from isolated islets from female NOD or B6 mice (8-12 weeks) to detect Gag gene messages. D: Cloning and sequencing of Gag gene PCR products in (C). The DNA sequences were aligned to a Gag gene cloned from Min6 insulinoma to identify ORFs. The numbers (n) of total sequenced clones for each of the mouse strains, NOD, B6, (NODxB6)F1 (F1), NOD.H2b (H2b) and B6.I-Ag7(Ag7) are shown, and the frequency of ORFs is calculated.

Article Snippet: The secondary antibodies used were anti-rat Ig-HRP (Cat#A10549, Invitrogen), AP-labelled secondary reagent for detecting rat IgG (AK5004, Vector Lab), and Avidin-HRP (Cat#RPN1231V, GE Healthcare).

Techniques: Staining, Labeling, Reverse Transcription Polymerase Chain Reaction, Isolation, Clone Assay, Sequencing

Journal: Immunology letters

Article Title: Endogenous Retrovirus Gag Antigen and Its Gene Variants Are Unique Autoantigens Expressed in the Pancreatic Islets of Non-obese Diabetic Mice

doi: 10.1016/j.imlet.2020.04.007

Figure Lengend Snippet: Identification of islet cell subset(s) expressing Gag antigen. A: Frozen sections of pancreas from female NOD mice were stained with biotin-labeled anti-CD105 antibody, followed by AP-Avidin as secondary and BCIP/NBT were used as substrate (left); or the section were stained with both biotin-labeled anti-CD105 and anti-Gag R187 antibodies, followed by with AP-Avidin and HRP-anti-rat IgG and then followed with BCIP/NBT and AEC substrates (right). B: Islet cell suspensions from female NOD mice (8-12 weeks) were stained with fluorescent-labeled antibodies shown. C: Cell sorting was performed using the stained cells in (B) to isolate CD45+ or CD45−/Sca-1+ islet cell subsets (left). RT-PCR was performed using total RNA (genomic DNA-free) from the sorted cells or total NOD islets to detect Gag gene expression.

Article Snippet: The secondary antibodies used were anti-rat Ig-HRP (Cat#A10549, Invitrogen), AP-labelled secondary reagent for detecting rat IgG (AK5004, Vector Lab), and Avidin-HRP (Cat#RPN1231V, GE Healthcare).

Techniques: Expressing, Staining, Labeling, Avidin-Biotin Assay, FACS, Reverse Transcription Polymerase Chain Reaction