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Western blot analysis of αA-crystallin-wt and its mutants, R12C, R21L, R21W, R49C, R54C, R116C and R116H expressed in HeLa cells.

By Ilangovan Raju (333566) and Edathara C. Abraham (91285)

Abstract

<p><i>A. Western blot analysis of HeLa cells individually expressed with αA-Crystallin:</i> Cells were individually transfected with YFP-tagged αA-wt and mutants. After 48 h transfection, cells were lysed and 5 µg of total protein was subjected to western blot. The blot was probed with human anti-αA antibody. The same blot was stripped and re-probed for β- actin to serves as a loading control. Nearly a similar level of expression of αA was evident in each of the transfected cells. The * indicates the non-speficific band. <i>B: Western blot analysis of HeLa cells co-expressed with CFP-tagged αA-wt and YFP-tagged αA-wt or its mutants</i>. Cells were co-transfected with YFP-tagged αA-wt and mutants with CFP-tagged αA-wt. After 48 h transfection, cells were lysed and 5 µg of total protein was subjected to western blot. The blot was probed with an antibody against human αA and the same blot was stripped and re-probed for actin. A similar level of expression of αA was detected in each group. <i>C: Western blot analysis of HeLa cells co-expressed with CFP-tagged αB-wt and YFP-tagged αA-wt or its mutants</i>. After 48 h transfection, 5 µg of total protein was subjected to western blot. The blot was probed with an antibody against human αA and the same blot was stripped and re-probed for anti-αB. The same blot was again stripped with anti-β-actin for loading control. The level of both αA and αB were nearly equal in each of the transfected cells.</p

Topics: Biochemistry, Medicine, Cell Biology, Genetics, blot, r116c, r116h, hela
Year: 2013
DOI identifier: 10.1371/journal.pone.0028085.g004
OAI identifier: oai:figshare.com:article/378837
Provided by: FigShare
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