A new assay for cytotoxic lymphocytes, based on a radioautographic readout of 111 In release, suitable for rapid, semi-automated assessment of limit-dilution cultures.

Détails

ID Serval
serval:BIB_A4470CCDED90
Type
Article: article d'un périodique ou d'un magazine.
Collection
Publications
Titre
A new assay for cytotoxic lymphocytes, based on a radioautographic readout of 111 In release, suitable for rapid, semi-automated assessment of limit-dilution cultures.
Périodique
Journal of Immunological Methods
Auteur(s)
Shortman K., Wilson A.
ISSN
0022-1759 (Print)
ISSN-L
0022-1759
Statut éditorial
Publié
Date de publication
1981
Volume
43
Numéro
2
Pages
135-152
Langue
anglais
Résumé
A new assay for cytotoxic T lymphocytes is described, of general application, but particularly suitable for rapid, semi-automated assessment of multiple microculture tests. Target cells are labelled with high efficiency and to high specific activity with the oxine chelate of 111 indium. After a 3-4 h incubation of test cells with 5 X 10(3) labelled target cells in V wells of microtitre trays, samples of the supernatant are spotted on paper (5 microliter) or transferred to soft-plastic U wells (25-50 microliter) and the 111 In release assessed by radio-autography. Overnight exposure of X-ray film with intensifying screens at -70 degrees C gives and image which is an intense dark spot for maximum release, a barely visible darkening with the low spontaneous release, and a definite positive with 10% specific lysis. The degree of film darkening, which can be quantitated by microdensitometry, shows a linear relationship with cytotoxic T lymphocyte dose up to the 40% lysis level. The labelling intensity and sensitivity can be adjusted over a wide range, allowing a single batch of the short half-life isotope to serve for 2 weeks. The 96 assays from a single tray are developed simultaneously on a single small sheet of film. Many trays can be processed together, and handling is rapid if 96-channel automatic pipettors are used. The method allows rapid visual scanning for positive and negative limit dilution cultures in cytotoxic T cell precursor frequency and specificity studies. In addition, in conjunction with an automated densitometer designed to scan microtitre trays, the method provides an efficient alternative to isotope counting in routine cytotoxic assays.
Mots-clé
Animals, Autoradiography, Cells, Cultured, Cytotoxicity Tests, Immunologic, Cytotoxicity, Immunologic, Dose-Response Relationship, Immunologic, Indium/diagnostic use, Mice, Oxyquinoline, Radioisotopes/diagnostic use, Solubility, T-Lymphocytes/immunology
Pubmed
Web of science
Création de la notice
10/04/2013 11:14
Dernière modification de la notice
03/03/2018 20:13
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