Objectives: A number of investigations have implicated a role for proteinases, especially metalloproteinases, in the etiology of endometriosis. The objectives of this study were to determine qualitative and quantitative differences in tissue proteinases obtained from human endometrium and peritoneum for comparison to samples from endometriotic lesions within the peritoneal cavity. Design: Recent studies from this laboratory, using zymographic methods, observed considerable differences between estrogen-dependent-metalloproteinases from rat uterus when analyzed by zymography in gels copolymerized with gelatin compared to transferrin. In the current study, a similar comparison of proteinases was made from the different human tissue samples by these two types of zymograms. Materials and Methods: Tissue samples were obtained from biopsies of human endometrium or from peritoneum and/or endometriotic lesions following hysterectomy or laparoscopy, homogenized in 0.1 M Triton X-100 and centrifuged. Samples of the supernatant were analyzed by zymography in SDS polyacrylamide gels copolymerized with either gelatin or denatured carboxymethylated transferrin. Results: There were significant differences in the metalloproteinases observed from endometrium when gelatin and transferrin zymograms were compared. Additionally, more patient to patient variabilities were observed in transferrin gel. For the most part, metalloproteinase from peritoneum was similar, both in regard to patient to patient and gelatin to transferrin comparisons. To this point, these enzymes are referred to as metalloproteinase in that they are inhibited by EDTA. In one patient, samples were obtained from uterine and ovarian peritoneum that were pathologically diagnosed as endometriotic. The metalloproteinases observed in these samples were present to a much lesser extent than those observed in previous peritoneal samples. The major proteolytic activities were observed as about 3 bands, around 45 kDa, only in gelatin gels and were not inhibited by EDTA. These proteinases were not observed in endometrial samples from this patient. Conclusions: Proteinases, especially metalloproteinases have been implicated in a number of pathologies, including endometriosis. We have demonstrated the presence of a number of metalloproteinase in human endometrium and uterine peritoneum by zymography. The involvement of one or more of these proteinases in endometriosis has not been established. However, we have demonstrated the presence of other types of proteinases in endometriotic lesions which are not inhibited by EDTA and not observed in endometrial samples.