Liu et al., 2017 - Zebrafish B Cell Development without a Pre-B Cell Stage, Revealed by CD79 Fluorescence Reporter Transgenes. Journal of immunology (Baltimore, Md. : 1950)   199(5):1706-1715 Full text @ J. Immunol.

Fig. 2

Predominant expression of CD79a and CD79b in kidney by in situ hybridization. (A) In situ hybridization of IgH-μ, CD79a, and CD79b antisense RNA probes to thin sections of zebrafish kidney. Images were processed with Nuance software to highlight hybridization signal stained by NBT/5-bromo-4-chloro-3-indolyphosphate. Right panels show background with sense probes. Scale bars, 400 μm. (B) RT-PCR analysis of CD79a and CD79b in zebrafish tissues. Image color is inverted for clarity.

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Stage: Adult

Fig. 3

GFP expression in CD79a and CD79b transgenic zebrafish identify B cells. (A) Diagram of production of transgenic constructs, made by long PCR from BACs containing CD79a and CD79b, where the first coding exon of each was replaced by an EGFP-PolyA segment. (B) Low power (left panels) and higher power (right panels) images of GFP expression in thin sections of kidney region of adult (4–6 mo) CD79a-GFP and CD79b-GFP zebrafish. Images were processed with Nuance software to reduce background autofluorescence. Scale bars, 100 μm (left), 200 μm (right). (C) Presence of CD79a:GFP+ and CD79b:GFP+ cells outside head kidney in adult fish. CD79a-GFP, ×4 cryosection. Scale bar, 100 μm (left), and original magnification ×10 image of cryosection. Scale bar, 200 μm (right). CD79a-GFP transgenic zebrafish also showing intestinal region. Scale bar, 200 μm. (D) Flow cytometry histogram plots of kidney marrow tissue from CD79a-GFP and CD79b-GFP lines. (E) GFP and GFP+ cell fractions were purified by electronic cell sorting from Lck-GFP, CD79a-GFP, and CD79b-GFP kidney tissue. Figure shows ethidium bromide staining of DNA amplified by RT-PCR from these samples. The lines indicate where parts of the image were joined.

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Stage: Adult

Fig. 4

Identification of developing and mature B cells in CD79a-GFP and CD79b-GFP × Rag2-mCherry double-transgenic zebrafish. (A) B cells in young CD79a-GFP and CD79b-GFP zebrafish larvae identified by coexpression with Rag2:mCherry, analyzed by flow cytometry. Wild-type zebrafish AB line is shown as a control. Representative data of four to seven sample analyses from each day postferilization zebrafish are shown. (B) Kidney marrow of 1–8 mo CD79a-GFP × Rag2-mCherry. Dotted region is GFPmCherry. (C) Five month CD79b-GFP × Rag2-mCherry zebrafish. Low-power (top) and high-power (bottom) images of thin sections of kidney and thymus are shown. Scale bars, 200 μm (top) and 100 μm (bottom). (D) Flow cytometry analysis of adult CD79b-GFP × Rag2-mCherry zebrafish kidney and thymus. Dotted region is GFPmCherry. Frequency of CD79b:GFP+ cells in kidney is 26.4%, in thymus is 5%. Data are representative of five separate CD79b/Rag2 transgenic fish kidney analyses.

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Stage: Adult

Fig. 6

Zebrafish B cell response to bacterial infection with CD79 reduction. (A) Low-power images of intestinal region of PBS control and bacteria injected CD79a-GFP fish at 1 wk postinfection. Scale bars, 1 mm. (B) Percentage of CD79a++ and CD79a+ cells in indicated tissues at 1, 2, and 4 wk postinfection; n = 3 each, mean ± SE. (C) Flow cytometry histograms of CD79a++ and CD79a+ cells in kidney and intestine tissue 1 wk postinfection, showing the clear increase in low CD79+ cells in intestine. (D) Increase in size of CD79a+ cells from intestine of infected fish, detected by flow cytometry using forward light scatter (FSC). (E) CD79a+ cells are enriched for secreted IgM compared with CD79a++ cells. RT-PCR analysis of β-actin, membrane IgH-μ (m), and secreted IgH-μ (s).

Acknowledgments:
This image is the copyrighted work of the attributed author or publisher, and ZFIN has permission only to display this image to its users. Additional permissions should be obtained from the applicable author or publisher of the image. Full text @ J. Immunol.