Validation Data

1. Body Weight and Spleen Weight Measurement

No significant differences in average body weight and spleen weight were detected between RenMab™ Mouse and wild type mouse.

2. Immune Cells Profiling

The percentages of immune cells in spleen were analyzed by flow cytometry. In RenMab™ Mouse, the percentage of B cells,T cells, NK cells, CD4+ T cells and CD8+ T cells in spleen were identical to those of wild type mice.

3. B Cell Development of RenMab™ Mouse

No significant differences in average body weight and spleen weight were detected between RenMab™ Mouse and wild type mouse.

IgM and IgD expression on B220+ splenocytes were analyzed by flow cytometry. Transitional type 1 (T1, B220+IgM+IgD), Transitional type 2 (T2, B220+IgM+IgD+), Mature (M, B220+IgMlowIgD) cell population.CD21 and CD23 expression on the surface of B220+ splenocytes were analyzed by flow cytometry. Marginal-zone (MZ, B220+CD21+CD23) and Follicular (FO, B220+CD21LOWCD23+) B cells were quantitatively analyzed.No significant difference was observed between RenMab™ mouse and wild type mouse.

4. Bone Marrow B Cell Development Analysis

B cell progenitor cells in bone marrow were analyzed by flow cytometry. Base on expression levels of B220 and CD43, B cell progenitor cells in bone marrow can be divided into 3 cell populations, B220lowCD43highIgMlow(Pro-B-cell), B220lowCD43intIgMlow(Pre-B-cell), and B220highCD43lowIgMhigh(Immature-B-cell). B220lowIgMIgDCD138 (Plasma cell ), B220+IgM+IgDCD38+ (Memory B cell ) in bone marrow or spleen were  also analyzed by flow cytometry. No significant difference was observed between RenMab™ mouse and wild type mouse.

5. Serum Ig Subtype Analysis

Different Ig subtypes in the serum of RenMab™ and WT(C57BL/6) mice were quantitatively measured by ELISA. No significant differences in IgA, IgG1, IgG2b, IgG2c, IgG3 and IgM levels were observed.

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