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. 2001 Aug 6;194(3):247–254. doi: 10.1084/jem.194.3.247

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© 2001 The Rockefeller University Press

This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/4.0/).

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Calpastatin expression does not inhibit IL-7 deprivation-induced apoptosis. (A) Long-term cultured pre-BI cells were transduced using pLZR-/IRES/GFP and pLZR-calpastatin/IRES/GFP. GFP⁺ cells were monitored and sorted. Pre-BI/mock, pre-BI/GFP, and pre-BI/calpastatin cells (10⁵ each) were collected and washed in ice-cold PBS. Cells were incubated in 30 μl of staining PBS with biotin-conjugated anti-B220, anti-IgM, or anti-CD43 antibody, washed, incubated with streptavidin-SPRD, and analyzed on a flow cytometer. (B) Pre-BI/mock, pre-BI/GFP, and pre-BI/calpastatin cells were cultured (0.25 × 10⁶ cells/ml) in medium with or without IL-7 for the apoptosis assay. Apoptosis was evaluated by staining cellular DNA. Values represent the mean of three independent experiments. Western blot (WB) analysis was performed using goat anti-calpastatin to confirm expression.

Calpastatin expression does not inhibit IL-7 deprivation-induced apoptosis. (A) Long-term cultured pre-BI cells were transduced using pLZR-/IRES/GFP and pLZR-calpastatin/IRES/GFP. GFP⁺ cells were monitored and sorted. Pre-BI/mock, pre-BI/GFP, and pre-BI/calpastatin cells (10⁵ each) were collected and washed in ice-cold PBS. Cells were incubated in 30 μl of staining PBS with biotin-conjugated anti-B220, anti-IgM, or anti-CD43 antibody, washed, incubated with streptavidin-SPRD, and analyzed on a flow cytometer. (B) Pre-BI/mock, pre-BI/GFP, and pre-BI/calpastatin cells were cultured (0.25 × 10⁶ cells/ml) in medium with or without IL-7 for the apoptosis assay. Apoptosis was evaluated by staining cellular DNA. Values represent the mean of three independent experiments. Western blot (WB) analysis was performed using goat anti-calpastatin to confirm expression.