A simplified and rapid staining method for the HeLa x skin fibroblast human hybrid cell neoplastic transformation assay

Marc Mendonca, R. J. Antoniono, C. Sun, J. L. Redpath

Research output: Contribution to journalArticle

34 Citations (Scopus)

Abstract

A simplified and rapid screening method for detecting radiation-induced neoplastically transformed foci in the HeLa x skin fibroblast human hybrid cell assay system has been developed. The method is based on the recent identification of the tumor-associated antigen in this system as intestinal alkaline phosphatase (IAP), and on the recent commercial development of a stable alkaline phosphatase chromogenic substrate solution, Western blue (WB). Cleavage of the substrate results in the production of a blue insoluble precipitate. It is shown that WB can be used on both viable and paraformaldehyde-fixed cells. Fixation does not noticeably reduce the IAP enzymatic activity. A direct comparison with the current method of immunoperoxidase (IMPO) staining indicates that the WB method is not only easier, but appears to be more sensitive in picking up weakly positive foci with a resulting higher (factor of 2.5) induced transformation frequency for 7 Gy of 137Cs γ radiation. Whereas the IMPO staining procedure is time- consuming and requires access to large amounts of expensive IAP-specific BD6 monoclonal antibody and peroxidase-labeled secondary antibody, the WB staining procedure is rapid and utilizes an inexpensive and readily available reagent. It should now allow this assay system to enter general use.

Original languageEnglish (US)
Pages (from-to)345-350
Number of pages6
JournalRadiation Research
Volume131
Issue number3
DOIs
StatePublished - 1992
Externally publishedYes

Fingerprint

phosphatases
Hybrid Cells
fibroblasts
staining
skin (animal)
rapid methods
alkaline phosphatase
Fibroblasts
Staining and Labeling
Skin
Alkaline Phosphatase
assays
cells
antibodies
antigens
radiation
reagents
Chromogenic Compounds
Radiation Dosage
cleavage

ASJC Scopus subject areas

  • Agricultural and Biological Sciences (miscellaneous)
  • Radiology Nuclear Medicine and imaging
  • Biophysics
  • Radiation

Cite this

A simplified and rapid staining method for the HeLa x skin fibroblast human hybrid cell neoplastic transformation assay. / Mendonca, Marc; Antoniono, R. J.; Sun, C.; Redpath, J. L.

In: Radiation Research, Vol. 131, No. 3, 1992, p. 345-350.

Research output: Contribution to journalArticle

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