This article updates our previous review of Ki67 published in Histopathology 10 years ago. In this period the numbers of papers published featuring this antibody has increased 10-fold from 338 to 3489 indicating the considerable enthusiasm with which this antibody has been studied. This review attempts to provide an update on the characterization of the Ki67 protein, its function and its use as a prognostic or diagnostic tool.

Monoclonal antibody Ki-67 is a reliable and easy means of accurately assessing the growth fraction of human neoplasms. Although the number of long-term follow-up studies is limited, it does appear to provide valuable prognostic information particularly in lymphoproliferative disease. Since the estimation of growth fraction is only one factor influencing tumour behaviour it would be naive to believe that measurement of this parameter alone, no matter how accurately, would provide the clinician with definitive prognostic information for all tumours. The antibody is also of use in research, providing a means of measuring proliferative activity in a variety of conditions besides malignancy, and may prove of value in monitoring tumour response to established and trial therapies.

For routine use, MIB1 or polyclonal Ki67 are the best proliferation markers in conventional histological preparations. The other markers tested cannot be recommended.

1 The ability of angiotensin II to modulate dopamine release from rat striatal slices in vitro and in the intact rat striatum in vivo was assessed by the microdialysis technique.2 In slices of rat striatum, angiotensin II (0.1-1.0 gM) induced a concentration-related increase in endogenous dopamine release which was maximal (approximately 250% above basal levels) within the first 2-4 min of agonist application and subsequently declined to near

The expression of cytokeratin intermediate filaments by a tumour has been accepted as evidence of an epithelial origin. Although there have been anecdotal reports of cytokeratin expression within tissues and neoplasms of non-epithelial origin, particularly muscle, there have been no comprehensive studies of its frequency and distribution. In order to investigate this we have studied 51 cases of normal smooth muscle and benign and malignant smooth muscle tumours using a panel of monoclonal antibodies against a range of intermediate filaments (cytokeratins, desmin and vimentin). Cytokeratin expression was noted overall in 50% of normal, benign and malignant smooth muscle tissues. Such expression tended to have a focal or patchy distribution. No case expressed cytokeratins in the absence of both desmin and vimentin. The implication of these findings for diagnostic immunocytochemistry is that intermediate filaments alone are not completely reliable markers of tumour histogenesis and should be used as part of a larger panel of monoclonal antibodies.

SUMMARY A series of 41 fresh and 36 routinely processed malignant melanomas were immunostained with a panel of 12 monoclonal antibodies reactive against a range of epithelial, lymphoid, and melanoma associated antigens. The aim of the study was to determine whether this panel of antibodies would be useful in diagnostically difficult cases for differentiating melanomas from other tumours, particularly carcinomas and lymphomas. The results confirmed that most unequivocal malignant melanomas can be identified by positivity for S100 protein and for the antigen recognised by antibody NK1/C3, and by negativity for epithelial and lymphoid antigens. The incidence of melanomas expressing cytokeratin antigens was higher, however, particularly in cryostat sections than has previously been reported. It is therefore suggested that a panel of antibodies with more than one marker in each category should be used for identifying melanomas in clinical practice.When the origin of a malignant tumour is unknown the two most likely alternative diagnoses are lymphoma and carcinoma. Recently, studies performed in several laboratories have established the patterns of antigenic expression characteristic of these two cate-

1 The ability of 5-HT4 (5-hydroxytryptamine4) receptor ligands to modify dopamine release from rat striatal slices in vitro and in the striatum of freely moving rats was assessed by the microdialysis technique. 2 The release of dopamine from slices of rat striatum continually perfused with Krebs buffer was enhanced by 5-HT4 receptor agonists; 5-HT (10 iM), 5-methoxytryptamine (5-MeOT; 10 gM), renzapride (10 gM) and (S)-zacopride (10 gM) maximally increased dopamine release by 133 + 5, 214 + 25, 232 + 29 and 264+69%, respectively (mean + s.e.mean, n =3-8). The drug-induced responses were maximal within the first 2 min of drug application, and subsequently declined. The non-selective 5-HT3/5-HT4 receptor antagonist, SDZ205-557 (10 gIM), failed to modify basal dopamine release from striatal slices but completely antagonized the (S)-zacopride (10 !LM)-induced increase in dopamine release. 3 To allow faster drug application, the modulation of dopamine release from rat striatal slices in a static release preparation was also investigated. The 5-HT4 receptor agonist, renzapride (10 /M) also enhanced dopamine release in this preparation (maximal increase = 214 + 35%, mean + s.e.mean, n = 14), whilst a lower concentration of renzapride (3 gM) was less effective. The renzapride-induced response was maximal within the first 2 min of drug application, before declining. In this preparation, the stimulation of dopamine release by renzapride (10 gM), was completely antagonized by the selective 5-HT4 receptor antagonist, GRI13808 (100 nM). In addition, both the Na+ channel blocker, tetrodotoxin (100 nM) and the non-selective protein kinase A inhibitor, H7 (100 nM) completely prevented the stimulation of dopamine release induced by renzapride (10 tM).4 In vivo microdialysis studies demonstrated that the 5-HT4 receptor agonists, 5-MeOT (10 gM), renzapride (100 gM) and (S)-zacopride (100 gM) maximally elevated extracellular levels of dopamine in the striatum by 220+20, 161+10 and 189+53%, respectively (mean+s.e.mean, n=5-9). A lower concentration of renzapride (10 gM) was less effective. The elevation of extracellular striatal dopamine levels induced by either renzapride (100 kM) or (S)-zacopride (100 kM) were completely antagonized by the non-selective 5-HT3/5-HT4 receptor antagonist, SDZ205-557 (100 jM). In addition, the elevation of extracellular levels of dopamine induced by either 5-MeOT (10 gM) or renzapride (100 gM) was completely prevented by the selective 5-HT4 receptor antagonist, GRI13808 (1 jM) and the renzapride (100 jgM)-induced response was also completely prevented by the non-selective protein kinase A inhibitor, H7 (1 gM). In this in vivo preparation, both GRI13808 (1 gM) and H7 (1 jM), when perfused alone, reduced extracellular levels of dopamine. 5 In conclusion, the present study provides evidence that the 5-HT4 receptor facilitates rat striatal dopamine release in vitro and in vivo.

Thirty-one cervical biopsies of invasive carcinoma have been studied by immunohistochemical means using the monoclonal antibody Ki67 to determine tumour cell proliferation rates. A wide range (10-50%) in the extent of Ki67 staining (expressed as the percentage of labelled tumour cells) was observed indicating considerable variation on tumour growth rates. There was no significant relationship between the percentage of positive cells and conventional histological parameters such as cell type or tumour differentiation. Immunostaining with monoclonal antibody Ki67 therefore provides a new approach to the assessment of cervical tumour biopsies which will require long term clinical follow-up to establish its prognostic significance. Images p179-a Figure 2