Generation of B cell–specific HSP90b1-deficient mice. (A) Genotype confirmation of KO and WT mice by PCR using allele-specific primers (flox indicates HSP90b1 gene floxed; del, HSP90b1 allele deleted; and cre, presence of cre gene). (B) Quantification of HSP90b1 in WT and KO splenic CD19⁺ cells by intracellular staining (open histogram indicates HSP90b1; shaded histogram, isotype control). (C) Immunoblot analysis of HSP90b1 using a HSP90b1 mAb in purified CD19⁺ splenocytes. (D) Quantification of HSP90b1 in different subset of B cells by intracellular staining (open histogram). (E) Comparative flow cytometric analysis of bone marrow cells for cell surface expression of B220 and λ5. Numbers on plots in panels D and E are the percentages of total cells in gate. (F) Phenotypic analysis of B220⁺ splenic B cells from KO and WT mice. Multiple experiments were conducted with similar findings. (G) Intracellular staining of HSP90b1 in WT and KO splenocytes. Various lineages were analyzed separately after gating with the indicated lineage-specific markers (open histogram indicates HSP90b1; shaded histogram, isotype control.