Methods of Antigen Recovery Vary in Their Usefulness in Unmasking Specific Antigens in Immunohistochemistry

Frost, Andra R.; Sparks, Dierick; Grizzle, William E.

doi:10.1097/00129039-200009000-00011

Cited by 49 publications

(40 citation statements)

References 23 publications

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“…[35][36][37] Specifically, sections (5 m thick) were cut from each block and mounted on Superfrost/Plus slides (Fisher Scientific, Pittsburgh, PA). All sections were cut 1 day prior to immunostaining, to avoid antigen decay, and were attached to the slide by heating in a 60°C oven for 1 hr.…”

Section: Immunohistochemical Stainingmentioning

confidence: 99%

“…[37][38][39] The slides were cooled and rinsed with deionized water. All sections were incubated with an aqueous solution of 3% hydrogen peroxide for 5 min and rinsed with Tris-buffered saline to reduce endogenous peroxidase activity, and then incubated with 1% goat serum for 1 hr at room temperature to reduce nonspecific background staining.…”

Section: Immunohistochemical Stainingmentioning

confidence: 99%

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Hormone receptors and proliferation in breast carcinomas of equivalent histologic grades in pre‐ and postmenopausal women

Talley

Grizzle

Waterbor

et al. 2001

Intl Journal of Cancer

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Breast cancers in younger, premenopausal women are more likely to exhibit an adverse prognostic profile (including negative steroid hormone receptors and a high rate of cellular proliferation) and poor outcome than breast cancers in postmenopausal women. It has been hypothesized that this adverse prognostic profile is a result of the higher histologic grade of breast cancers in pre-compared with post-menopausal women. To assess the association of expression of steroid hormone receptors and indicators of proliferation while controlling for histologic grade, we identified 100 infiltrating ductal carcinomas from premenopausal women 45 years of age or younger and 100 from postmenopausal women 65 years of age or older. The carcinomas were selected so that the histologic grades (low versus high) were distributed equally between the 2 groups. Estrogen receptors (ER), progesterone receptors (PR), p27 Kip1 and Ki-67 (to measure rate of proliferation) were assessed by immunohistochemistry and compared between groups. Clinical information and survival data were also analyzed. ER content was lower and proliferation was higher in carcinomas in premenopausal women (p ‫؍‬ 0.048 and p ‫؍‬ 0.005, respectively). By univariate analysis, p27 Kip1 and PR were not different between the groups; however, in multivariate analysis, p27 Kip1 was higher in postmenopausal women, but only in a subgroup with highly proliferative carcinomas. Overall survival was similar in the pre-and postmenopausal women. Furthermore, low p27 Kip1 and African-American ethnicity predicted a poorer overall survival in the premenopausal, but not in the postmenopausal, women in our study. After controlling for histologic grade, a lower expression of ER and a higher proliferative index were detected in breast carcinomas in premenopausal women. Therefore, some prognostic indicators, such as ER and proliferative rate, may be more closely associated with menopausal status than histologic grade. Our data also suggest that some prognostic factors are not equally effective as predictors of survival in pre-and postmenopausal women. Prognosis in women with breast cancer has been reported to differ according to age and menopausal status. Studies examining this issue have varied in design, age categorization and outcome measures. A few large population-based studies and more limited analyses 1-5 have indicated that women under the ages of 30 -40 years have a worse prognosis. Some retrospective studies have attempted to explain the less favorable outcome in younger women and have reported that breast cancers in younger women exhibit more adverse characteristics, including a high clinical stage at diagnosis, high histologic grade, lack of steroid hormone receptors and increased proliferation. 6 -9 Multivariate analyses to assess associations of age and menopausal status with other clinical and pathologic characteristics have not consistently retained either variable as an independent adverse prognosticator, 8 although several multivariate analyses have indicated that young ag...

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Section: Immunohistochemical Stainingmentioning

confidence: 99%

Section: Immunohistochemical Stainingmentioning

confidence: 99%

Hormone receptors and proliferation in breast carcinomas of equivalent histologic grades in pre‐ and postmenopausal women

Talley

Grizzle

Waterbor

et al. 2001

Intl Journal of Cancer

Self Cite

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“…The most frequently used immunomarkers for animals in the routine diagnosis are polyclonal, as they contain antibodies to a wider range of epitopes, increasing sensitivity of the method, sometimes in detriment of specificity. Antigen retrieval techniques, especially the heat-induced ones, have been widely used in human pathology since the 80's, allowing reliable immunodetection of epitopes distorted by formalin fixation (4,17,21) . These techniques have proven efficiency in routine veterinary pathology, permitting the utilization of a wider spectrum of markers, including monoclonal and those intended for human use, as shown by our results.…”

Section: Discussionmentioning

confidence: 99%

“…To overcome this drawback, antibodies which present cross reactivity with human and animal antigens have been applied. Other limitations of the method are represented by antigen masking by formalin fixation, reducing specific antigen-antibody binding (4,17,21) . In this respect, an extraordinary progress has been achieved with the production of a wide variety of monoclonal antibodies to epitopes resistant to formalin fixation, thanks to the hybridoma technology, with the use of antigen retrieval methods and powerful amplifying systems.…”

Section: Introductionmentioning

confidence: 99%

Immunohistochemistry in diagnostic veterinary pathology: a critical review

Ruiz¹,

Alessi²,

Chagas³

et al. 2005

J. Bras. Patol. Med. Lab.

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key words unitermos abstractImmunohistochemistry has become a practical and widely used tool for diagnosis in human pathology since the 70's. However, its application in veterinary diagnostic pathology has not been so common, especially due to the lack of specific antibodies. To overcome this drawback, antibodies which present cross reactivity with human and animal antigens have been applied. The purpose of the present study was to test the cross reactivity of some antibodies intended for the human pathology, which may be used in animal tissues, with the help of antigen retrieval and amplification systems. In the present study it was confirmed that many of the antibodies produced for use in human histopathology might be applied in veterinary pathology. Further studies are needed to increase the list of applicability of these antibodies to different animal species. It must be stressed that in this type of study some variables, such as clone of antibody, dilution, antigen retrieval method, and detection system, have to be evaluated. Lab • v. 41 • n. 4 • p. 263-70 • agosto 2005 a more accurate diagnosis. This fact has brought to the attention of veterinary pathologists the need to improve the application of immunohistochemistry in their daily routine, following the tendency of human diagnostic pathology. Immunohistochemistry

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“…[1][2][3] Recent innovations in antigen-retrieval procedures for formalin-fixed paraffin embedded tissue sections have brought increased reliability to immunostaining of routinely produced paraffin-embedded tissue sections. [4][5][6][7][8][9] This is now an established diagnostic tool for general use in the histopathology laboratory and in our institute, Toyama Medical and Pharmaceutical University Hospital, 486 out of 3854 cases (13%) examined in 2002 required immunostaining in addition to routine H&E staining. Because around 10 antibodies were applied per case on average, almost 5000 immunostained specimens were produced to assist in diagnosis.…”

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confidence: 99%

Improved 1-h rapid immunostaining method using intermittent microwave irradiation: practicability based on 5 years application in Toyama Medical and Pharmaceutical University Hospital

et al. 2004

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Immunostaining depending on antigen-antibody specificity is the commonest approach for determining the localization of specific antigens in tissue sections. This procedure is applicable not only with frozen or specially fixed samples, but also has proved reliable with formalin-fixed paraffin-embedded tissue sections through improvement of antigen-retrieval. Immunostaining is thus firmly established as a tool for diagnostic pathology and in our institute multiple antibodies are applied for 13-15% of the cases examined, as well as H&E staining. With the standard approach, approximately 3 h is necessary from the beginning of deparaffinization till covering sections with the Envision system. We utilized intermittent microwave irradiation for 10 min during hybridization with primary and secondary antibodies in a special moist-chamber, to achieve all immunostaining steps within 1 h in 178 primary antibodies frequently used for diagnostic pathology. According to our 5 years experience, such microwave irradiation not only obtained significant specific staining for enhancing the specificity of antigen-antibody reactions, but also inhibited nonspecific binding. We present herein the details of the methodology and recommendations for its application with particular primary antibodies. This method can contribute to savings in time and energy, allowing pathologists to rapidly obtain diagnostic information.

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Methods of Antigen Recovery Vary in Their Usefulness in Unmasking Specific Antigens in Immunohistochemistry

Cited by 49 publications

References 23 publications

Hormone receptors and proliferation in breast carcinomas of equivalent histologic grades in pre‐ and postmenopausal women

Hormone receptors and proliferation in breast carcinomas of equivalent histologic grades in pre‐ and postmenopausal women

Immunohistochemistry in diagnostic veterinary pathology: a critical review

Improved 1-h rapid immunostaining method using intermittent microwave irradiation: practicability based on 5 years application in Toyama Medical and Pharmaceutical University Hospital

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