Optimization of proliferating cell nuclear antigen (PCNA) immunohistochemical staining: A comparison of methods using three commercial antibodies, various fixation times, and antigen retrieval solution

K. Mintze, N. Macon, K. E. Gould, George Sandusky

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Abstract

The purpose of this study was to identify the optimal staining for cell proliferation in paraffin embedded sections of rabbit and pig ileum. Methods comparing zinc formalin or 10% buffered formalin, 3 PCNA antibodies (19A2, PC10, MAB424), fixation time intervals at 4, 8, 24, and 48 hr, and antigen retrieval solution were developed. Staining of nuclei in rabbit ileum was optimal after 4-8 hr in zinc formalin with the PC10, 19A2, and MAB424 antibodies, without antigen retrieval. All 3 antibodies produced optimal staining of nuclei in pig ileum fixed in zinc formalin 4-8 hr without antigen retrieval. After 4 hr fixation in 10% neutral buffered formalin (NBF), nuclei in both pig and rabbit were lightly stained in comparison to those fixed in zinc formalin. Minimal or no staining was seen at 8, 24, and 48 hr fixation in 10% NBF for rabbit ileum. Weak staining was seen at 28 and 48 hr in 10% NBF for pig ileal nuclei. Antigen retrieval methods did enhance nuclear staining with zinc formalin fixation and 10% NBF in the pig but not in the rabbit. The PC10 and 19A2 antibodies stained more intensely than the MAB424 antibody.

Original languageEnglish (US)
Pages (from-to)25-30
Number of pages6
JournalJournal of Histotechnology
Volume18
Issue number1
StatePublished - 1995
Externally publishedYes

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Proliferating Cell Nuclear Antigen
Formaldehyde
Staining and Labeling
Antigens
Antibodies
Zinc
Swine
Ileum
Rabbits
Paraffin
Cell Proliferation

Keywords

  • antigen retrieval solution
  • fixation
  • proliferating cell nuclear antigen

ASJC Scopus subject areas

  • Anatomy
  • Cell Biology

Cite this

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title = "Optimization of proliferating cell nuclear antigen (PCNA) immunohistochemical staining: A comparison of methods using three commercial antibodies, various fixation times, and antigen retrieval solution",
abstract = "The purpose of this study was to identify the optimal staining for cell proliferation in paraffin embedded sections of rabbit and pig ileum. Methods comparing zinc formalin or 10{\%} buffered formalin, 3 PCNA antibodies (19A2, PC10, MAB424), fixation time intervals at 4, 8, 24, and 48 hr, and antigen retrieval solution were developed. Staining of nuclei in rabbit ileum was optimal after 4-8 hr in zinc formalin with the PC10, 19A2, and MAB424 antibodies, without antigen retrieval. All 3 antibodies produced optimal staining of nuclei in pig ileum fixed in zinc formalin 4-8 hr without antigen retrieval. After 4 hr fixation in 10{\%} neutral buffered formalin (NBF), nuclei in both pig and rabbit were lightly stained in comparison to those fixed in zinc formalin. Minimal or no staining was seen at 8, 24, and 48 hr fixation in 10{\%} NBF for rabbit ileum. Weak staining was seen at 28 and 48 hr in 10{\%} NBF for pig ileal nuclei. Antigen retrieval methods did enhance nuclear staining with zinc formalin fixation and 10{\%} NBF in the pig but not in the rabbit. The PC10 and 19A2 antibodies stained more intensely than the MAB424 antibody.",
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author = "K. Mintze and N. Macon and Gould, {K. E.} and George Sandusky",
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T2 - A comparison of methods using three commercial antibodies, various fixation times, and antigen retrieval solution

AU - Mintze, K.

AU - Macon, N.

AU - Gould, K. E.

AU - Sandusky, George

PY - 1995

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N2 - The purpose of this study was to identify the optimal staining for cell proliferation in paraffin embedded sections of rabbit and pig ileum. Methods comparing zinc formalin or 10% buffered formalin, 3 PCNA antibodies (19A2, PC10, MAB424), fixation time intervals at 4, 8, 24, and 48 hr, and antigen retrieval solution were developed. Staining of nuclei in rabbit ileum was optimal after 4-8 hr in zinc formalin with the PC10, 19A2, and MAB424 antibodies, without antigen retrieval. All 3 antibodies produced optimal staining of nuclei in pig ileum fixed in zinc formalin 4-8 hr without antigen retrieval. After 4 hr fixation in 10% neutral buffered formalin (NBF), nuclei in both pig and rabbit were lightly stained in comparison to those fixed in zinc formalin. Minimal or no staining was seen at 8, 24, and 48 hr fixation in 10% NBF for rabbit ileum. Weak staining was seen at 28 and 48 hr in 10% NBF for pig ileal nuclei. Antigen retrieval methods did enhance nuclear staining with zinc formalin fixation and 10% NBF in the pig but not in the rabbit. The PC10 and 19A2 antibodies stained more intensely than the MAB424 antibody.

AB - The purpose of this study was to identify the optimal staining for cell proliferation in paraffin embedded sections of rabbit and pig ileum. Methods comparing zinc formalin or 10% buffered formalin, 3 PCNA antibodies (19A2, PC10, MAB424), fixation time intervals at 4, 8, 24, and 48 hr, and antigen retrieval solution were developed. Staining of nuclei in rabbit ileum was optimal after 4-8 hr in zinc formalin with the PC10, 19A2, and MAB424 antibodies, without antigen retrieval. All 3 antibodies produced optimal staining of nuclei in pig ileum fixed in zinc formalin 4-8 hr without antigen retrieval. After 4 hr fixation in 10% neutral buffered formalin (NBF), nuclei in both pig and rabbit were lightly stained in comparison to those fixed in zinc formalin. Minimal or no staining was seen at 8, 24, and 48 hr fixation in 10% NBF for rabbit ileum. Weak staining was seen at 28 and 48 hr in 10% NBF for pig ileal nuclei. Antigen retrieval methods did enhance nuclear staining with zinc formalin fixation and 10% NBF in the pig but not in the rabbit. The PC10 and 19A2 antibodies stained more intensely than the MAB424 antibody.

KW - antigen retrieval solution

KW - fixation

KW - proliferating cell nuclear antigen

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