anti mouse  (Bioss)

Journal: bioRxiv

Article Title: Rare SH2B3 coding variants identified in lupus patients impair B cell tolerance and predispose to autoimmunity

doi: 10.1101/2023.04.27.538529

Figure Lengend Snippet: A-C . IgG immune-complex deposits in mice treated with PBS or pristane. ( A ) Representative immunofluorescence images showing IgG (green) and podocin (red) staining in the kidney sections of pristane/PBS-treated Sh2b3 E372K mice 20 weeks following treatment with ( B ) pristane and ( C ) PBS and numbers of kidney sections that are positive and negative for IgG ICs in the glomeruli of Sh2b3 E372K mice treated with ( B ) pristane and ( C ) PBS. D-E. Numbers of kidney sections with indicated glomerular score 20 weeks following treatment with ( D ) pristane and ( E ) PBS. G-R . Cellular and serological phenotypes of SW HEL -mHEL 3× chimeric mice. Frequencies of non-HEL-specific BM ( F ) mature and ( G ) immature B cells, ( H ) HEL-specific Pro-B cells, non-HEL-specific splenic ( H ) transitional and ( I ) mature B cells, relative surface IgM expression on HEL-specific splenic ( J ) transitional, ( L ) FO and ( M ) MZ B cells in WT (black circles) or mHEL 3× (red circles) recipients receiving BM from SW HEL - Sh2b3 +/+ (unfilled), SW HEL - Sh2b3 E372K/+ (yellow filling) or SW HEL - Sh2b3 E372K/E372K (orange filling) donors. Levels of anti-HEL ( N ) IgM and ( O ) IgG in wildtype and mHEL 3× mice receiving BM from SW HEL - Sh2b3 +/+ , - Sh2b3 E372K/+ and - Sh2b3 E372K/E372K donors measured as OD 405-605 by ELISA. Frequencies of ( P ) non-HEL-specific ABCs, ( Q ) HEL-specific and ( R ) non-HEL-specific CD21 - CD23 - mature B cells as percentages of splenic lymphocytes. ( S ) Percentages of apoptotic BM immature B cells in untreated, 20 ng/mL IL-4 only, 5 μg/mL anti-IgM (α-IgM) only, and 5 μg/mL α-IgM + 20 ng/mL IL-4 conditions. BAFF-R surface expression on ( T ) transitional and ( U ) mature B cells measured by flow cytometry as median fluorescence intensity (MFI). Data in A-C is pooled from two independent experiments. Results in D-U are representative of 2-3 independent experiments. Fisher’s exact test was used for the statistical analyses in B-E. Means in D-U are shown as bars, two-way ANOVA was used for statistical analysis in G-U. Significance levels for two-way ANOVAs are indicated with asterisks. *: p < 0.05, **: p < 0.01, ***: p < 0.001, ****: p < 0.0001.

Article Snippet: OCT-embedded, cryopreserved mouse kidneys were cut into 7-μm sections, fixed with acetone, blocked with blocking buffer (1× PBS, 3% BSA, 0.5% Triton X-100) and stained with goat anti-mouse Podocin antibody (Santa Cruz Biotechnology; cat # sc-22296) ( Supplementary Table 8 ) and then with Alexa Fluor 594-conjugated donkey anti-goat IgG (H+L) and Alexa Fluor 488-conjugated donkey anti-mouse IgG (H+L) antibodies (Invitrogen; cat # A-21202).

Techniques: Immunofluorescence, Staining, Expressing, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Fluorescence

Journal: bioRxiv

Article Title: Dynein-mediated trafficking and degradation of nephrin in diabetic podocytopathy

doi: 10.1101/2022.10.01.510475

Figure Lengend Snippet: The histological and ultrastructural features of diabetic glomerulopathy and podocytopathy developed in mice three months after receiving i.p. injection of s treptozotocin, as shown by periodic acid-Schiff staining and Masson Trichrome staining (a, Scale Bars = 50µm) and transmission electron microscopy (b,Scale Bars = 1µm). In mice with STZ-mediated DN, podocytopathy presented with effacement (black arrows) of foot processes, or cortical dislocation of slit diaphragms (white arrows). Podocytes with STZ-induced diabetic injury are characterized by the dislocation of nephrin and podocin from the slit diaphragm to the cytosol (arrows) as detected by immunogold labeling (c). Hyperglycemia (d) and albuminuria (e), quantified as the albumin/creatinine ratio, in mice with STZ-mediated DN ( *p<0 . 05 vs . pre injection, n=6). Histology and ultrastructural features of diabetic glomerulopathy were expressed as the average glomerular area (f, n=6 mice x 5 glomeruli per mouse) and the percentage of glomerular capillaries covered by effaced foot processes (g, n=6). *p<0 . 05 vs . vehicle control.

Article Snippet: For immunogold labeling , 8 nm sections were placed on carbon-coated and glow-discharged, formvar-coated nickel slot grids; blocked grids were incubated in a mouse anti-podocin antibody (Sigma Aldrich, #SAB4200810) or a rabbit anti-nephrin antibody (Invitrogen, # PA5-91907) at room temperature, followed by a goat anti-rabbit Au 12 nm conjugated (Jackson ImmunoResearch Laboratories Inc. #111-205-144) and a goat anti-mouse Au 6 nm conjugated (Jackson ImmunoResearch Laboratories Inc. #115-195-146).

Techniques: Injection, Staining, Transmission Assay, Electron Microscopy, Labeling