Skip to main content
NCBI home page
Journal of Clinical Pathology logoLink to Journal of Clinical Pathology
. 1994 May;47(5):448–452. doi: 10.1136/jcp.47.5.448

Microwave antigen retrieval in immunocytochemistry: a study of 80 antibodies.

E C Cuevas 1, A C Bateman 1, B S Wilkins 1, P A Johnson 1, J H Williams 1, A H Lee 1, D B Jones 1, D H Wright 1
PMCID: PMC502024  PMID: 7517960

Abstract

AIMS--To evaluate the effect of microwave irradiation on the staining quality of a range of commonly used primary antibodies in archival, formalin fixed, paraffin wax embedded material, with emphasis on antibodies that have previously worked successfully only on frozen tissue. METHODS--Immunocytochemistry (streptavidin-biotin complex technique) was performed on histological sections of a range of normal and pathological tissues, after varying treatment with microwave irradiation. The staining quality of each antibody was compared with that achieved without prior treatment of the sections or after enzyme predigestion. RESULTS--Microwave irradiation permitted successful immunostaining with 20 antibodies that stained only frozen tissues before. The staining characteristics of 21 antibodies that were already known to stain formalin fixed, paraffin wax embedded material were improved. Another 39 antibodies did not show enhanced staining with microwave irradiation. The method preserves tissue morphology and produces more consistent staining than that achieved by enzyme predigestion with many antibodies. Microwave irradiation may also allow some primary antibodies to be used at higher working dilutions. The citrate buffer used in this study avoids the necessity of exposure to heavy metal salts. CONCLUSIONS--Microwave antigen retrieval represents an important technical advance within immunocytochemistry that will greatly increase the range of antibodies which can be used to study formalin fixed, paraffin wax embedded tissues.

Full text

PDF
448

Images in this article

449Image
on p.449
450Image
on p.450
451Image
on p.451

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Anstee D. J., Gardner B., Spring F. A., Holmes C. H., Simpson K. L., Parsons S. F., Mallinson G., Yousaf S. M., Judson P. A. New monoclonal antibodies in CD44 and CD58: their use to quantify CD44 and CD58 on normal human erythrocytes and to compare the distribution of CD44 and CD58 in human tissues. Immunology. 1991 Oct;74(2):197–205. [PMC free article] [PubMed] [Google Scholar]
  2. Cattoretti G., Becker M. H., Key G., Duchrow M., Schlüter C., Galle J., Gerdes J. Monoclonal antibodies against recombinant parts of the Ki-67 antigen (MIB 1 and MIB 3) detect proliferating cells in microwave-processed formalin-fixed paraffin sections. J Pathol. 1992 Dec;168(4):357–363. doi: 10.1002/path.1711680404. [DOI] [PubMed] [Google Scholar]
  3. Cattoretti G., Pileri S., Parravicini C., Becker M. H., Poggi S., Bifulco C., Key G., D'Amato L., Sabattini E., Feudale E. Antigen unmasking on formalin-fixed, paraffin-embedded tissue sections. J Pathol. 1993 Oct;171(2):83–98. doi: 10.1002/path.1711710205. [DOI] [PubMed] [Google Scholar]
  4. Chiu K. Y. Use of microwaves for rapid immunoperoxidase staining of paraffin sections. Med Lab Sci. 1987 Jan;44(1):3–5. [PubMed] [Google Scholar]
  5. Cuevas E., Jones D. B., Wright D. H. Immunohistochemical detection of tumour growth fraction (Ki-67 antigen) in formalin-fixed and routinely processed tissues. J Pathol. 1993 Apr;169(4):477–478. doi: 10.1002/path.1711690415. [DOI] [PubMed] [Google Scholar]
  6. Estrada J. C., Brinn N. T., Bossen E. H. A rapid method of staining ultrathin sections for surgical pathology TEM with the use of the microwave oven. Am J Clin Pathol. 1985 May;83(5):639–641. doi: 10.1093/ajcp/83.5.639. [DOI] [PubMed] [Google Scholar]
  7. Garrido M. C., Cordell J. L., Becker M. H., Key G., Gerdes J., Jones M., Gatter K. C., Mason D. Y. Monoclonal antibody JC1: new reagent for studying cell proliferation. J Clin Pathol. 1992 Oct;45(10):860–865. doi: 10.1136/jcp.45.10.860. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Gerdes J., Becker M. H., Key G., Cattoretti G. Immunohistological detection of tumour growth fraction (Ki-67 antigen) in formalin-fixed and routinely processed tissues. J Pathol. 1992 Sep;168(1):85–86. doi: 10.1002/path.1711680114. [DOI] [PubMed] [Google Scholar]
  9. Gerdes J., Lemke H., Baisch H., Wacker H. H., Schwab U., Stein H. Cell cycle analysis of a cell proliferation-associated human nuclear antigen defined by the monoclonal antibody Ki-67. J Immunol. 1984 Oct;133(4):1710–1715. [PubMed] [Google Scholar]
  10. Hsu S. M., Raine L., Fanger H. Use of avidin-biotin-peroxidase complex (ABC) in immunoperoxidase techniques: a comparison between ABC and unlabeled antibody (PAP) procedures. J Histochem Cytochem. 1981 Apr;29(4):577–580. doi: 10.1177/29.4.6166661. [DOI] [PubMed] [Google Scholar]
  11. Login G. R., Dvorak A. M. Microwave energy fixation for electron microscopy. Am J Pathol. 1985 Aug;120(2):230–243. [PMC free article] [PubMed] [Google Scholar]
  12. Mason D. Y., Cordell J. L., Gaulard P., Tse A. G., Brown M. H. Immunohistological detection of human cytotoxic/suppressor T cells using antibodies to a CD8 peptide sequence. J Clin Pathol. 1992 Dec;45(12):1084–1088. doi: 10.1136/jcp.45.12.1084. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Mayers C. P. Histological fixation by microwave heating. J Clin Pathol. 1970 Apr;23(3):273–275. doi: 10.1136/jcp.23.3.273. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Mepham B. L., Frater W., Mitchell B. S. The use of proteolytic enzymes to improve immunoglobulin staining by the PAP technique. Histochem J. 1979 May;11(3):345–357. doi: 10.1007/BF01005033. [DOI] [PubMed] [Google Scholar]
  15. Merz H., Rickers O., Schrimel S., Orscheschek K., Feller A. C. Constant detection of surface and cytoplasmic immunoglobulin heavy and light chain expression in formalin-fixed and paraffin-embedded material. J Pathol. 1993 Jul;170(3):257–264. doi: 10.1002/path.1711700307. [DOI] [PubMed] [Google Scholar]
  16. Sawhney N., Hall P. A. Ki67--structure, function, and new antibodies. J Pathol. 1992 Oct;168(2):161–162. doi: 10.1002/path.1711680202. [DOI] [PubMed] [Google Scholar]
  17. Shi S. R., Key M. E., Kalra K. L. Antigen retrieval in formalin-fixed, paraffin-embedded tissues: an enhancement method for immunohistochemical staining based on microwave oven heating of tissue sections. J Histochem Cytochem. 1991 Jun;39(6):741–748. doi: 10.1177/39.6.1709656. [DOI] [PubMed] [Google Scholar]
  18. Shimoyama Y., Hirohashi S. Expression of E- and P-cadherin in gastric carcinomas. Cancer Res. 1991 Apr 15;51(8):2185–2192. [PubMed] [Google Scholar]

Articles from Journal of Clinical Pathology are provided here courtesy of BMJ Publishing Group

RESOURCES