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Identification of molecular components of the centrosphere in the mitotic spindle of sea urchin eggs
Monoclonal antibodies were prepared to identify molecular components specific to the mitotic apparatus of sea urchin eggs. The mitotic apparatus or asters induced within unfertilized eggs by taxol treatment were isolated from Strongylocentrotus purpuratus and used for immunization of mice. After fus...
Formato: | Texto |
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Lenguaje: | English |
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The Rockefeller University Press
1985
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2113690/ https://www.ncbi.nlm.nih.gov/pubmed/3894377 |
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collection | PubMed |
description | Monoclonal antibodies were prepared to identify molecular components specific to the mitotic apparatus of sea urchin eggs. The mitotic apparatus or asters induced within unfertilized eggs by taxol treatment were isolated from Strongylocentrotus purpuratus and used for immunization of mice. After fusion with spleen cells, the supernatant of hybridomas were screened in two stages by indirect immunofluorescence staining, first on isolated sea urchin mitotic spindles in 96-well microtiter plates to identify rapidly potential positive hybridomas, and second, on whole mitotic eggs on coverslips to distinguish between spindle-specific staining and adventitious contamination. Two hybridomas, SU4 and SU5, secreted antibodies reactive to microtubule-containing structures in eggs during the course of development. They preferentially stained the centrosphere both in isolated mitotic apparatus and in whole metaphase eggs, which was further confirmed by staining the isolated centrospheres with these antibodies. SU4 recognized a major 190-kD polypeptide on immunoblots as well as a species at 180 and 20 kD, whereas hybridoma SU5 stained a species at 50 kD. Thus, these polypeptides may be components of the centrosphere. |
format | Text |
id | pubmed-2113690 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 1985 |
publisher | The Rockefeller University Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-21136902008-05-01 Identification of molecular components of the centrosphere in the mitotic spindle of sea urchin eggs J Cell Biol Articles Monoclonal antibodies were prepared to identify molecular components specific to the mitotic apparatus of sea urchin eggs. The mitotic apparatus or asters induced within unfertilized eggs by taxol treatment were isolated from Strongylocentrotus purpuratus and used for immunization of mice. After fusion with spleen cells, the supernatant of hybridomas were screened in two stages by indirect immunofluorescence staining, first on isolated sea urchin mitotic spindles in 96-well microtiter plates to identify rapidly potential positive hybridomas, and second, on whole mitotic eggs on coverslips to distinguish between spindle-specific staining and adventitious contamination. Two hybridomas, SU4 and SU5, secreted antibodies reactive to microtubule-containing structures in eggs during the course of development. They preferentially stained the centrosphere both in isolated mitotic apparatus and in whole metaphase eggs, which was further confirmed by staining the isolated centrospheres with these antibodies. SU4 recognized a major 190-kD polypeptide on immunoblots as well as a species at 180 and 20 kD, whereas hybridoma SU5 stained a species at 50 kD. Thus, these polypeptides may be components of the centrosphere. The Rockefeller University Press 1985-08-01 /pmc/articles/PMC2113690/ /pubmed/3894377 Text en 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/). |
spellingShingle | Articles Identification of molecular components of the centrosphere in the mitotic spindle of sea urchin eggs |
title | Identification of molecular components of the centrosphere in the mitotic spindle of sea urchin eggs |
title_full | Identification of molecular components of the centrosphere in the mitotic spindle of sea urchin eggs |
title_fullStr | Identification of molecular components of the centrosphere in the mitotic spindle of sea urchin eggs |
title_full_unstemmed | Identification of molecular components of the centrosphere in the mitotic spindle of sea urchin eggs |
title_short | Identification of molecular components of the centrosphere in the mitotic spindle of sea urchin eggs |
title_sort | identification of molecular components of the centrosphere in the mitotic spindle of sea urchin eggs |
topic | Articles |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2113690/ https://www.ncbi.nlm.nih.gov/pubmed/3894377 |