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Parameters of the labeling of mitogen-activated murine lymphocytes by [35S]methionine for two-dimensional gel electrophoresis. I. Effect of culture conditions.

Abstract：

:Labeling with [35S]methionine at a high specific activity is essential to the facile preparation of 2-dimensional gel electrophoretograms with the analytical 2-dimensional charge-size separation procedure (Anderson's ISODALT system). Mitogen-activated T and B lymphocytes subjected to low methionine concentrations would not proceed through cell cycle. In the case of activated B lymphocytes, the use of fetal bovine serum (FBS), dialyzed to lower endogenous methionine concentrations, prevented B cell growth even in the presence of otherwise satisfactory levels of methionine. High concentrations of [35S]methionine (greater than 300 mCi/1) induced B cell death, apparently by radiation damage. Despite these problems, good radioautograms and radiofluorograms of 2D electrophoretograms could be prepared by labeling activated B or T cells in bulk (10(6) cells/ml) with high specific activity [35S]methionine. The polypeptides labeled may be a biased sample since lymphoid cells do not proceed through cell cycle under these conditions. Small numbers (10(3] of activated T cells also yielded satisfactory samples but labeling of small numbers of activated B cells was not possible.

journal_name

J Immunol Methods

journal_title

Journal of immunological methods

authors

Kettman JR,Kuhn L,Young P,Lefkovits I

doi

10.1016/0022-1759(86)90051-7

subject

Has Abstract

pub_date

1986-04-03 00:00:00

pages

53-64

issue

1

eissn

0022-1759

issn

1872-7905

pii

0022-1759(86)90051-7

journal_volume

88

pub_type

杂志文章

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