Estimation of hormone receptor status in fine-needle aspirates and paraffin-embedded sections from breast cancer using the novel rabbit monoclonal antibodies SP1 and SP2

We describe a method of immunocytochemical assessment of estrogen receptor (ER) status on alcohol-fixed smears obtained by fine-needle aspiration (FNA) from breast cancer patients, using a commercially available rabbit monoclonal antibody anti-ER (SP1) without any antigen retrieval. A series of 40 aspirates were analyzed and the results of ER status were compared with the respective formalin-fixed tissue using the same procedure and with assessment by the classical method using the mouse monoclonal antibody 6F11 (anti-ER) with antigen retrieval on paraffin sections. Twenty-four out of the 40 cases examined were positive at least by two methods and 16 were negative for all three determinations. The results obtained in the ER immunocytochemical assay on aspirates and paraffin sections using the antibody SP1 and those obtained on paraffin sections using the antibody 6F11 were quite similar. In one case the material was insufficient to interpret the reaction in the cytological specimen and only one case, with focal positivity reaction on paraffin sections, was negative in the cytological specimen. The intensity of nuclei staining in cytological smears of breast cancer cells was stronger than that observed by traditional methods. We also assessed progesterone receptor (PR) status on 40 paraffin-sections from breast cancer patients, using a commercially available rabbit monoclonal antibody anti-PR (SP2), with the same characteristics described for anti-ER (SP1). The results were compared with assessment by the classic method with mouse monoclonal antibody 1A6 (PR) on paraffin sections and total agreement was observed. Of the 40 cases examined, 18 were positive and 22 were negative for the two determinations. We conclude that the application of the ER method on alcohol-fixed smears/paraffin sections with the rabbit monoclonal antibody SP1, and the PR method on paraffin sections with the rabbit monoclonal antibody SP2, provide several advantages, such as high sensitivity and specificity of the reaction, stronger immunostaining, shorter procedures times, and avoidance of antigenic retrieval methods.     

Methodology of immunohistological detection of oestrogen receptor in human breast carcinoma in formalin-fixed, paraffin-embedded tissue: a comparison with frozen section methodology

We describe a method of immunohistochemically assessing oestrogen receptor status on routinely processed formalinfixed tissue, using a commercially available monoclonal antibody (Abbott H222). with pronase predigestion of tissue sections and overnight antibody incubation. The staining was assessed using the H score system. A series of 94 cases of breast cancer were analysed and the results were compared with assessment by oestrogen receptor immunocytochemical assay performed on frozen section. Direct comparison of the paired sets of H scores obtained with frozen tissue and formalin-fixed tissue showed a highly significant correlation of 0.8 ( P < 0.001) between the two methods of oestrogen receptor assessment. Chi-squared analysis using H score cut off points of 50 and 100 also showed a similar significant association ( P < 0.001). We conclude that this oestrogen receptor method, applicable to formalin-fixed, paraffin-embedded tissue, gives accurate results on routinely fixed tissue and could be used as an alternative to other methods.     

Potential value of estrogen receptor immunocytochemical assay in formalin-fixed breast tumors.

Sixty-two primary breast carcinomas were analyzed for estrogen receptor (ER) by both the dextran-coated charcoal (DCC) technique and estrogen receptor immunocytochemical assay (ER-ICA) on cryostat and permanent sections. Paraffin sections of formalin-fixed breast tissue underwent DNase pretreatment to expose the nuclear antigenic site as described by P. Shintaku and J. H. Said (Am J Clin Pathol 87:161, 1987). The results of immunocytochemical staining agreed with those of the DCC biochemical assay in 89% of paraffin-sectioned tissue and in 94% of the cryostat sections. Comparison of the results of ER-ICA on permanent and frozen sections showed 85% agreement (kappa statistic = 0.704). This study suggests that ER can be demonstrated immunocytochemically on paraffin-sectioned breast tissue. However, although highly specific, immunoperoxidase determination on paraffin-embedded tissue is less sensitive than that on frozen tissue. The commercial source of DNase, length of incubation, and tissue fixation are important factors in the demonstration of ER immunoreactivity. The assay may offer an alternative for assessment of ER when tissue is not suitable or available for biochemical assay or conventional cytochemical analysis.     

Immunocytochemical evaluation of estrogen receptor on archival Papanicolaou-stained fine-needle aspirate smears

The availability of limited fine-needle aspirate smears necessitates the selection of immunocytochemical (IC) methods that allow reuse of Pap-stained smears to assess the estrogen receptor (ER) status of breast carcinoma. The objective of the current study was to compare IC evaluation of ER status on FNA smears by three methods: 1) ER-ICA using H222 monoclonal antibody performed on slides fixed in formaldehyde-methanol-acetone; 2) destained Pap slides using 1D5 antibody; and 3) Pap-stained slides without destaining using the same 1D5 antibody. Two representative Pap smears of breast carcinoma were selected from 48 cases of breast carcinoma in which ER was previously evaluated by ER-ICA. One of these Pap smears was used as such and the other was destained prior to immunostaining by a modified ABC method using 1D5 monoclonal antibody. The number of cells with positive nuclear staining was expressed as a percentage and the intensity of staining was semiquantitatively scored on a scale of 1+ to 3+. The degree of agreement between the three methods was evaluated statistically by weighted kappa statistics. Thirty cases (63%) showed varying degrees of positive staining while 18 cases (38%) were entirely negative by all three methods. Significant discrepancies in the number of cells with positive staining and in the intensity of staining between the three methods occurred in 40% and 23% of the cases and was mainly due to a reduction in the number of cells with positive staining and the intensity of staining using Pap slides in comparison to ER-ICA. Weighted kappa agreement of the percentage of cells with positive staining using Pap-stained slides and destained Pap-slides in comparison to ER-ICA was 0.75 and 0.64, respectively, and that for the intensity of staining was 0.75 and 0.66, respectively. Therefore, IC evaluation of ER using Pap-stained smears as such or destained Pap smears compared favorably with ER-ICA. However, Pap-stained smears used as such for ER immunostaining showed a slightly better agreement with ER-ICA than destained Pap smears. Because significant differences in ER-IC staining can occur with any of the immunocytochemical methods, a negative result is less reliable as an indicator of true ER status than a positive result.     

Evaluation of estrogen receptor immunocytochemical assay (ER-ICA) and estrogen receptor enzyme immunoassay (ER-EIA) for the determination of estrogen receptor status in breast carcinoma. A comparative study on frozen sections, cytological smears and tissue homogenates.

One hundred thirty three cases of primary breast cancer were analyzed for estrogen receptors (ERs) by two different techniques: monoclonal ER immunocytochemical assay (ER-ICA) and ER enzyme immunoassay (ER-EIA). The overall concordance between ER-ICA and ER-EIA was 89.5% despite the semiquantitative nature of the immunocytochemical evaluations. There was a significant correlation between these two methods in linear regression analysis. The correlations were evident both for ER detection on frozen sections as well, as on fine needle aspirates. Correlations between histoscores calculated independently by three pathologists might overcome the objections of subjectivity of ER-ICA evaluations. The results suggest that ER-ICA can be useful in assessing ERs in breast cancer, especially in small primary tumours, primary inoperable breast cancer, recurrences and metastases.     

Whither cytosolic estrogen receptor assays? A comparison of commercially available kits for estrogen receptor assay

Frozen tissue sections and cytosols from 89 specimens of breast and ovarian tumours have been assayed for the presence of estrogen receptor (ER) or related protein using four commercially available monoclonal antibody methods. These were estrogen receptor enzyme immunoassay (ER-EIA), estrogen receptor enzyme immunocytochemical assay (ER-ICA), ER D5 antigen immunoradiometric assay and ER D5 antigen immunocytochemical assay. The results have been compared with those obtained using a standard dextran coated charcoal steroid binding assay (ER-DCC). The correlation coefficient (r) between ER-DCC and ER-EIA results was 0.72 while that of both monoclonal antibody cytosol methods and their respective immunocytochemical assays was 0.66. ER-ICA gave additional valuable information concerning receptor heterogeneity in breast cancer sections. However, the correlation between ER D5 antigen assays and both ER-DCC and ER-EIA was weak (r less than 0.4). We conclude that there are a number of methodological advantages in using the kit systems including their ability to detect receptor presence in small tumour specimens (e.g., "Tru-cut" biopsies) but that their usefulness is limited by the current lack of widely available monoclonal based methods for the concurrent determination of progestogen receptor. We believe that, once these are available, immunocytochemical technology could offer an alternative method of determining the steroid receptor concentration in both ovarian and breast tumours, thus obviating the need for costly and time-consuming cytosolic methods, with their inherent difficulties of quality control.     

Automated immunohistochemical estrogen receptor in fixed embedded breast carcinomas.

The authors immunohistochemically assessed the presence of estrogen receptor (ER) in formalin-fixed, paraffin-embedded tissue sections of 68 breast carcinomas by an automated method using Pronase (CalBiochem, La Jolla, CA) predigestion and alkaline phosphatase detection (Method 1). These results were compared with those obtained by an automated peroxidase-antiperoxidase method with DNAse pretreatment of fixed embedded sections (Method 2), with ER immunostain on frozen sections (Method 3), and with biochemical results (dextran-coated charcoal cytosolic [DCC] assay). Compared with the DCC assay, Methods 1, 2, and 3 gave sensitivities of 54%, 25%, and 89%, respectively. The sensitivity for Method 1 was increased to 74% in those cases with DCC results showing greater than 50 fmol/mg protein. These findings indicate that ER immunohistochemical studies on formalin-fixed paraffin-embedded tissues (as assayed by Method 1) provide useful clinical information when the results are positive. A negative result, especially if surrounding normal elements are not positive, may indicate no receptors, receptor levels less than 50 fmol/mg protein, or improper tissue preservation. In the absence of fresh tissue for ER assay by DCC assay or of frozen sections for immunostaining, and with an understanding of its limitations, this method may be useful.     

Automated immunohistochemical estrogen receptor in fixed embedded breast carcinomas. Comparison with manual immunohistochemistry on frozen tissues

Estrogen receptor (ER) status of breast carcinomas determines prognosis and treatment. Biochemical ER assays are expensive and time-consuming and require fresh tumor. Immunohistochemical ER was assessed in 68 breast carcinomas, by an automated method using routinely processed formalin-fixed paraffin-embedded tissues, and manually with the use of snap-frozen tissues with a monoclonal anti-ER and peroxidase-antiperoxidase technique. The paraffin sections were digested with DNase to enhance development of signal. Positive nuclear ER was obtained in 9 (13%) fixed tissues and 36 (53%) frozen tissues. The sensitivity, specificity, and predictive value of a positive test result, as compared with the biochemical assay, were 25%, 100%, and 100% for the paraffin section technique, and 89%, 88%, and 89% for the frozen sections. Although it is specific, lack of sensitivity, resulting from loss of ER with fixation and room temperature handling, renders this immunohistochemical technique unacceptable on fixed tissues. However, ER immunostain on frozen tissue is an acceptable alternative to biochemical assay.     

Oestrogen receptor staining of paraffin-embedded breast carcinomas following short fixation in formalin: a comparison with cytosolic and frozen section receptor analyses

This paper describes an improved immunohistochemical method for demonstrating oestrogen receptor (OR) protein in paraffin-embedded sections of tissue fixed for 1.5 h in formalin. Thirty-two cases of infiltrating ductal breast carcinoma were stained with a monoclonal anti-OR antibody (H222), using a standard streptavidin-biotin method, following pretreatment with pronase. OR counts in paraffin sections were compared with those of frozen sections and with cytosolic values determined by a dextran-coated charcoal method. Twenty-seven of the carcinomas were OR-positive in paraffin sections. There was concordance between the paraffin section and the frozen section-determined receptor status in 30 cases (94 per cent) and a strong correlation was observed (r = 0.76; P less than 0.0001). Similarly, OR counts in paraffin sections correlated with cytosolic OR values (r = 0.60; P less than 0.001) and there was concordance in 97 per cent of cases. The percentage of positively-stained tumour cells in paraffin sections ranged from 0 to 94 per cent with staining intensities comparable to those seen in frozen sections. Staining of paraffin sections identified more OR-positive tumours than either frozen section staining or cytosolic assay. This study validates immunohistochemical OR analysis in formalin-fixed, paraffin-embedded breast carcinomas using a commercial anti-OR antibody.     

Immunostaining of estrogen receptor,progesterone receptor,MIB1 antigen,and c-erbB-2 oncoprotein in cytologic specimens: A simplified method with formalin fixation

An effective but simple fixation protocol for the immunocytochemical staining of cytologic smears for estrogen and progesterone receptors, the Ki-67 antigen (using MIB1 antibody), andc-erbB-2protein is described. One hundred twenty-seven smears from a variety of malignant and benign breast lesions showed good preservation of antigenicity when subjected to the following fixation protocol: Freshly made smears were air-dried for 20 min to 14 h at 22°C before immersing in 10% buffered formalin for 2–14 h. Immunostaining followed microwave-stimulated epitope retrieval. There was strong concordance of staining with corresponding tissue sections in 15 cases of malignant tumors (ER: r = 0.7381; PR: r = 0.6684; MIB1: r = 0.7234). Immunostaining staining, when delayed for 5–10 days in about half the smears, showed no noticeable difference in reactivity, attesting to effective storage of the formalin-fixed smears at room temperature. Diagn. Cytopathol. 17:127–133, 1997. © 1997 Wiley-Liss, Inc.     

Immunohistochemistry of estrogen receptor protein in paraffin sections. Effects of enzymatic pretreatment and cobalt chloride intensification

Monoclonal antibodies provide important tools for the demonstration of estrogen receptors (ERs) in cases of breast cancer. This study reports an improved immunohistochemical method for the demonstration of ER in formalin-fixed paraffin-embedded tissue samples using the Abbott monoclonal antibody to ER protein. Tissue sections were pretreated briefly with trypsin, followed by DNase before the performance of the immunohistochemical reaction and cobalt chloride was used to intensify the color of the diaminobenzidine reaction product. In 20 cases, the results in paraffin sections were compared with biochemical assays with the dextran-coated charcoal technique or with immunohistochemistry performed on frozen sections. There was an excellent correlation between the results obtained with all three methods. The introduction of cobalt chloride into the chromogen solution significantly increased the sensitivity of this approach as compared with the use of diaminobenzidine alone.     

Fine-needle aspiration biopsy of malignant melanoma: a cytologic and immunocytochemical analysis

The immunoreactivity of alcohol-fixed cell blocks from 15 fine-needle aspiration (FNA) specimens of malignant melanoma was investigated using monoclonal antibodies to keratin and vimentin intermediate filaments, melanoma cytoplasmic antigen (HMB-45), and S-100, as well as polyclonal antibodies to S-100. The results were compared with the immunoprofiles obtained using formalin-fixed surgical specimens from 10 of the same patients. In all cases, immunostaining for keratin was negative and immunostaining for vimentin was positive. Immunostaining for HMB-45 was positive in 13/15 aspirates and in 9/10 surgical specimens. Immunostaining for S-100 protein showed the greatest variability in staining, with 5/15 fine needle aspiration biopsies and 9/10 surgical specimens being positive using the polyclonal antibody and only 1/15 FNA specimens and 7/10 surgical specimens being positive using the monoclonal S-100 reagent. Our findings indicate that immunocytochemical studies can be very useful as an adjunct in the FNA diagnosis of melanoma. Also included in our series is an unusual variant of malignant melanoma, the so-called signet ring melanoma. Given the location of the anal verge, the use of immunocytochemical markers was essential in establishing the correct diagnosis in this case. While S-100 protein is of limited value as a marker of melanoma in alcohol-fixed FNA specimens, a definitive diagnosis of malignant melanoma can be made using a panel of antibodies including keratin, vimentin, and HMB-45.     

Utility of the paraffin-embedded section method on the detection of estrogen receptor from breast cancer tissues--comparison of the paraffin-embedded section method (6F11 and 1D5) with frozen section (H222) and dextran-coated charcoal (DCC) ones]

From March 1988 to December 1990, we detected estrogen receptor (ER) from breast cancer tissues with the dextran-coated charcoal (DCC) method. From September 1990 to March 1998, we repeated the above with the frozen section method (cloneH222; DAINABOT). With the paraffin-embedded section method using two antibodies of anti-ER (clone6F11; NOVOCASTRA and clone 1D5; MBL), we examined the ER of the same 185 primary breast cancer tissues. We had already detected these tissues with the frozen section method, and we also applied the same procedure for the 43 primary breast cancer tissues which had already been detected with the DCC method. We compared these data. The positive rates of ER with DCC, H222, 6F11 and 1D5 were 49%, 53%, 53% and 52% respectively, which were within the reported range. The accuracy between H222 and 6F11 which was calculated as the percentage that were in positive and negative concordance by the two methods, was 88%. The accuracies between H222 and 1D5, between 6F11 and 1D5, between DCC and 6F11, between DCC and 1D5 were respectively 89%, 96%, 79%, and 77%. Although the accuracy between DCC and the paraffin-embedded section was not necessarily high, we obtained a higher concordance between the frozen and paraffin-embedded sections. The highest concordance existed between 6F11 and 1D5. The 30% of negative cases with DCC were positive with paraffin-embedded section. Among these methods of ER detection from breast cancer tissues, the paraffin-embedded section method seemed to be the most useful.     

Immunocytochemical staining of progesterone receptor in paraffin sections of human breast cancers.

Monoclonal antibodies (mAB) against progesterone receptor (PR) and the peroxidase-antiperoxidase (PAP) method to visualize PR in paraffin sections from 68 human breast cancers were used. Ten mAB, which recognize human PR on frozen sections, were tested. Six could detect PR in paraffin sections, with Li 417, LET 456, and LET 126 giving the best results. LET 126 antibody was used for most further studies. The effects of fixation with picric acid-formaldehyde (PAF), buffered-formalin, or with Bouin solution were investigated; all fixation methods allowed PR immunolabeling, although PAF or buffered formalin usually gave the best results. Positive staining was seen in the nucleus of carcinoma cells. Variations in intensity and extent of immunoreactivity were observed in all sections and among different regions of the same specimen. These were probably related to the heterogeneity of the tumor cell population. Results were compared with the PR content in the respective tumor tissues, determined by steroid-binding assay, and with immunocytochemistry on frozen sections. It was shown that there were correlations between the immunocytochemical staining (positive or negative) and the steroid binding assay (80%) and between the immunocytochemical staining on paraffin sections and on frozen sections (78%). Weaker intensity and fewer number of PR-positive cells were found for paraffin-embedded tumors. Estrogen receptors were also detected on adjacent sections from the same paraffin-embedded tissues by use of monoclonal anti-ER antibodies (ERICA-kit[Abbott Laboratories, Chicago, IL]) and DNase pretreatment. In conclusion, this immunocytochemical method for detection of PR and ER on paraffin sections offers an alternative to frozen tissue. It allows histologic and immunocytochemical studies on the same sample and retrospective studies on stored tissue blocks.     

Demonstration of oestrogen receptors in paraffin wax sections of breast carcinoma using the monoclonal antibody 1D5 and microwave oven processing.

This study aimed at assessing the usefulness of a new monoclonal antibody (1D5) for the demonstration of oestrogen receptors (ER) in paraffin wax sections, using brief microwave processing rather than proteolytic predigestion. Routinely processed paraffin wax sections of 50 cases of breast carcinoma with known ER concentrations, estimated by the standard dextran-coated charcoal (DCC) biochemical assay, were examined using the avidin-biotin complex-immunoperoxidase technique. The results were assessed semiquantitatively, using a five grade scoring system. Of the 50 cases examined, 37 were positive and six were negative by both DCC and immunohistology. Of the remaining seven cases, three (6%) were negative by DCC but positive with immunohistology, and four (8%) were positive with DCC and negative with immunohistology. The DCC results of the latter four cases were 10, 14, 14 and 16 fmol/mg protein which is at the lowest level of positivity, our cutoff point being less than 10 fmol. The monoclonal antibody 1D5, as used in this study, can provide easily assessed reliable information about the ER status of breast carcinoma using routinely processed paraffin wax sections.     

Use of monoclonal antibody for assessment of estrogen receptor content in fine-needle aspiration biopsy specimen from patients with breast cancer

With increasing emphasis on application of fine-needle aspiration biopsy (FNAB) for an earlier detection of breast cancer, there is clearly a need for a method that requires only a small number of tumor cells for hormone receptor analysis. An immunocytochemical assay utilizing monoclonal antiestrophilin antibody and the peroxidase-antiperoxidase technique has been shown to be highly specific and sensitive for the detection of estrogen receptor (ER) in breast cancer tissues. To assess the usefulness of this technique in FNAB, 62 cases of primary, recurrent, and metastatic breast cancers were studied. The findings from the aspirated cells employing estrogen receptor immunocytochemical assay (ER-ICA) were compared with results obtained from cell population of the same tumors following their removal using both cytochemical (ER-ICA) and biochemical (dextran-coated charcoal assay) methods for ER determination. Overall, there was a 92% concordance between biochemical and cytochemical results. This result suggests that anti-ER monoclonal antibody in immunocytochemical analysis is an effective tool in assessment of ER content in breast cancers. The technique can be easily performed at community hospitals and is well suited for specimens of insufficient size for biochemical assay. It may extend the scope of FNAB and make ER studies possible on material unsuitable for biochemical assay from sites other than breast.     

Estrogen receptor analysis on biopsies and fine-needle aspirates from human breast carcinoma. Correlation of biochemical and immunohistochemical methods using monoclonal antireceptor antibodies.

Monoclonal antibodies against estrogen receptor (ER) were used for determination of ER status immunocytochemically in histologic specimens from 192 primary breast carcinomas. All tumors were also assayed biochemically for ER with the dextran-coated charcoal method (DCC). The comparison of biochemically and immunocytochemically determined ER status showed concordant results in 80% (P less than 0.0001). In only 2 cases (1%) with low ER levels (less than 20 fmol/mg protein) immunocytochemistry failed to detect ER. ER positivity determined with a semiquantified approach based on intensity and heterogeneity of immunocytochemical staining correlated significantly with biochemically determined ER levels (P = 0.0001). In a series of fine-needle aspirates of 34 breast carcinomas sufficient cell material was available for ER immunocytochemistry (ER-ICA). Overall, the results of ER-ICA in fine-needle aspirates were concordant with ER-ICA in histologic specimens in 88% of the samples. In a few cases with weak positivity of ER-ICA in histologic specimens, ER-ICA was negative in fine-needle aspirates. In no case was there a false-positive immunocytochemical ER determination in a tumor aspirate. Thus, ER-ICA seems to be a reliable assay which can be performed in histologic and cytologic specimens.     

Detection of estrogen receptors with monoclonal antibodies in routinely processed formalin-fixed paraffin sections of breast carcinoma. Use of DNase pretreatment to enhance sensitivity of the reaction

This study describes an improved immunohistochemical method for the sensitive and specific identification of estrogen receptors (ERs) in paraffin sections from formalin-fixed and routinely processed breast carcinoma tissues, using DNase pretreatment to expose nuclear antigenic sites and commercially available immunoreagents (including monoclonal antibody) in kit form. Results were compared with dextran-coated charcoal cytosolic assay (DCC) and with conventional immunohistochemistry on frozen sections. Sensitivity and specificity for determinations on paraffin sections were 88% and 86%, respectively, and statistical analysis showed very good agreement between DCC and paraffin sections (kappa = 0.805). The DNase technic on paraffin sections allows excellent correlation between histologic characteristics and ER status and reduces DCC sampling error resulting from stromal dilution and tumor variability. This method offers a reliable and reproducible alternative when tissue is not suitable or unavailable for DCC or frozen tissue analysis and can be used for retrospective studies on stored tissue blocks.     

An evaluation of potentially suitable fixatives for immunoperoxidase staining of estrogen receptors in imprints and frozen sections of breast carcinoma

The estrogen receptor (ER) content of breast carcinoma is generally accepted as valuable in predicting clinical outcome and tumour response to hormonal manipulation. We applied a new immunocytochemical assay for estrogen receptors (Abbott ERICA Monoclonal) to 20 breast tumours, and examined the efficacy of 16 fixation procedures before immunoperoxidase staining of frozen sections and imprint preparations. Our findings indicate that the fixatives of choice are periodatelysine-paraformaldehyde at 22 degrees C, or 10% formalin followed by acetone at -10 degrees C. These fixation procedures are simpler, less time-consuming, and provide superior staining, tumour cytomorphology and higher ER values than the 3-reagent sequence recommended by Abbott Laboratories. There was a significant correlation between the ER scores in the frozen sections and the imprints. Positive ER cytosol results correlated with the staining index in the frozen sections, and the ER scores in the imprints. We conclude that imprints are suitable preparations for ER analysis by the immunoperoxidase technique, particularly for small tumour specimens.     

Demonstration of progesterone receptors in paraffin wax sections of breast carcinoma.

Several immunohistological methods for the demonstration of progesterone receptors were tried on routinely processed paraffin wax sections of breast carcinoma, using Abbott's PgR-ICA monoclonal antibody. The best results were obtained with the avidin-biotin-immunoperoxidase complex method with no prior trypsinisation or DNAse digestion, and with imidazole added to the final diaminobenzidine developing solution. A simple semiquantitative scoring system was used to assess the staining results which were then compared with the results obtained by a standard dextran-coated charcoal biochemical assay. Of 31 cases examined, the results of the two methods were concordant in 25 (81%) of cases. This is near the higher end of the concordance range obtained by several other authors using frozen sections. The discordance encountered in a few cases was possibly the result of sampling errors which are more likely to occur with the chemical rather than the histological method. It is concluded that the method described here is fairly reliable and would greatly simplify the process of assessment of progesterone receptors in breast, and possibly other tumours.