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Detection of endometriosis with the use of plasma protein profiling by surface-enhanced laser desorption/ionization time-of-flight mass spectrometry

2007; Elsevier BV; Volume: 87; Issue: 4 Linguagem: Inglês

10.1016/j.fertnstert.2006.08.095

1556-5653

Haiyuan Liu, Jinghe Lang, Qunfang Zhou, Dan Shan, Qizhai Li,

nanoparticles nucleation surface interactions

In this prospective case-control study, we determined the role of surface-enhanced laser desorption and ionization time-of-flight mass spectrometry in the detection of histologically proven endometriosis. In the plasma of the group with endometriosis, there were 20 different protein peaks, and the classifier showed a sensitivity of 87.5% and a specificity of 80% in the diagnosis of endometriosis. In this prospective case-control study, we determined the role of surface-enhanced laser desorption and ionization time-of-flight mass spectrometry in the detection of histologically proven endometriosis. In the plasma of the group with endometriosis, there were 20 different protein peaks, and the classifier showed a sensitivity of 87.5% and a specificity of 80% in the diagnosis of endometriosis. Endometriosis is a common gynecological disease characterized by the growth of endometrial tissue outside the uterus, causing infertility and pelvic pain. The diagnosis of endometriosis is based on the histological identification of endometrial glands and stroma through invasive procedures such as laparoscopic surgery (1The American Fertility SocietyRevised American Fertility Society classification of endometriosis: 1985.Fertil Steril. 1985; 43: 351-352Abstract Full Text PDF PubMed Google Scholar). Up to now, there has been no reliable noninvasive method for detection of the disease, even though serum CA125 level has been widely used as a serological marker of endometriosis. In fact, the serum CA125 level is more useful in excluding epithelial cancer following the disease and in detecting recurrence (2Bast Jr, R.C. Xu F.J. Yu Y.H. Barnhill S. Zhang Z. Mill G.B. CA125: the past and the future.Int J Biol Markers. 1998; 13: 179-187Crossref PubMed Scopus (366) Google Scholar, 3Chen F.P. Soong Y.K. Lee N. Lo S.K. The use of serum CA125 as a marker for endometriosis in patients with dysmenorrhea for monitoring therapy and for recurrence of endometriosis.Acta Obstet Gynecol Scand. 1998; 77: 665-670Crossref PubMed Scopus (55) Google Scholar) than as a diagnostic method. A noninvasive and simple diagnostic method is urgently needed in both clinical practice and basic research.Recently developed proteomic techniques, such as surface-enhanced laser desorption and ionization time-of-flight mass spectrometry (SELDI-TOF MS), have been introduced for biomarker discovery. The SELDI-TOF MS system consists of a protein chip, mass analyzer, and bioinformatics software (4Chapman K. The Protein Chip Biomarker System from Ciphergen Biosystems: a novel proteomics platform for rapid biomarker discovery and validation.Biochem Soc Trans. 2002; 30: 82-87Crossref PubMed Google Scholar). The protein chips contain spots with different types of chromatographic material, i.e., hydrophobic, hydrophilic, ion-exchange, and immobilized metallic ion matrices. Small amounts of protein combine directly with the biochip by affinity capture, and are then analyzed by TOF MS to generate protein-profiling spectra. Surface-enhanced laser desorption and ionization enable a direct analysis of human body fluid with high speed, high throughput, and easy control. The new approach was successfully used in distinguishing the serum of healthy individuals from the serum of those with malignant diseases of the ovary (5Petricoin E.F. Ardekani A.M. Hitt B.A. Levine P.J. Fusaro V.A. Steinberg S.M. et al.Use of proteomic patterns in serum to identify ovarian cancer.Lancet. 2002; 359: 572-577Abstract Full Text Full Text PDF PubMed Scopus (2837) Google Scholar), prostate gland (6Qu Y.S. Adam B.L. Yutaka Y. Ward M.D. Cazares L.H. Schellhammer P.F. et al.Boosted decision tree analysis of surface-enhanced laser desorption/ionization mass spectral serum profiles discriminates prostate cancer from noncancer patients.Clin Chem. 2002; 48: 1835-1843PubMed Google Scholar), bladder (7Vlahou A. Schellhammer P.F. Mendrinos S. Patel K. Kondylis F.I. Gong L. et al.Development of a novel proteomic approach for the detection of transitional cell carcinoma of the bladder in urine.Am J Pathol. 2001; 158: 1491-1501Abstract Full Text Full Text PDF PubMed Scopus (393) Google Scholar), liver (8Poon T.C. Yip T.T. Chan A.T. Yip C. Yip V. Mok T.S. et al.Comprehensive proteomic profiling identifies serum proteomic signatures for detection of hepatocellular carcinoma and its subtypes.Clin Chem. 2003; 49: 752-760Crossref PubMed Scopus (233) Google Scholar), and lung (9Xiao X. Liu D. Tang Y. Guo F. Xia L. Liu J. et al.Development of proteomic patterns for detection lung cancer.Dis Markers. 2003; 19: 33-39Crossref PubMed Scopus (89) Google Scholar). The aim of this study was to determine the utility of plasma proteomic profiling with the use of SELDI-TOF MS for the detection of endometriosis.Eighty-seven patients who had been clinically diagnosed with endometriosis before surgery were enrolled in the study. All participants gave informed consent. All plasma samples were preoperatively collected. After surgery, 52 patients were histologically diagnosed with endometriosis, and their plasma samples were used for SELDI-TOF MS analysis. These patients were of reproductive age (range, 24–46 years), and had normal menstrual cycles. The staging of endometriosis was determined according to the revised scoring system of the American Fertility Society (1The American Fertility SocietyRevised American Fertility Society classification of endometriosis: 1985.Fertil Steril. 1985; 43: 351-352Abstract Full Text PDF PubMed Google Scholar). Plasma samples were taken from 46 female, age-matched blood donors from the blood transfusion department of Peking Union Medical College Hospital, Beijing, China. The blood donors were normal according to pelvic examination, vaginal sonography, and serum CA125 level. This study was approved by the Ethics Committee of Peking Union Medical College.All samples were obtained by venipuncture, and drawn into vacutainers with 3.2% sodium citrate and centrifuged at 3,000 rpm for 8 minutes at room temperature. The plasma was aliquoted and stored at −80°C until use.Plasma samples were thawed on ice and centrifuged at 10,000 rpm for 10 minutes. Twenty microliters of U9 buffer (9 M urea, 2% 3-[3-(Cholamidopropyl)dimethylammonio]-1-proanesulfonate (CHAPS), and 50 mM Tris-HCl, pH 9) were added to 10 μL of each plasma sample, and the solutions were vortexed for 30 minutes. The resulting plasma solutions were further diluted by 360 μL of buffer solution (50 mM potassium acetate, pH 4.0). Protein chips (WAX2, Ciphergen Biosystems, Freemont, CA) were preequilibrated with 200 μL of buffer solution twice. One hundred microliters of diluted plasma sample were added to each spot of WAX2 protein chip in a bioprocessor (Ciphergen Biosystems). The bioprocessor was then sealed and agitated on a platform shaker for 60 minutes.After discarding the remaining samples, the chips were washed with 200 μL of buffer solution for 5 minutes. This procedure was repeated three times, with two brief water rinses between applications of buffer solution. The chips were removed from the bioprocessor and air-dried. Each spot on the chip received two applications of 0.5 μL of a saturated solution of α-cyano-4-hydroxycinnamic acid in 50% v/v acetontrile and 5% v/v trifluoroacetic acid, and the chips were air-dried.All chips were analyzed with the use of the protein biological system II (PBS II) Protein Chip Reader (Ciphergen Biosystems). Data were collected by an average of 150 laser shots, with a detector sensitivity of 8 and a laser intensity of 195. The highest mass for detection was 50,000 Daltons (Da), with an optimization range between 2,000–20,000 Da. Mass accuracy was calibrated using the All-in-One Peptide Molecular Mass Standard (Ciphergen Biosystems).The SELDI-TOF mass spectra were obtained and analyzed using Ciphergen Protein Chip Software 3.1.1 (Ciphergen Biosystems). All spectra were compiled, and peak intensities were normalized in a mass range between 2,000–20,000 Da. The noise was filtrated using signal-to-noise ratio. An automatic peak detection program was used.Thirty-six samples from the endometriosis (EM) group and 32 samples from the control group were randomly selected as the training set. The remaining 16 EM samples and 14 control samples were designated as the masked set. The training set was used to compare the peak intensities of protein profiling between the two groups, and to set up a classification and regression tree as a diagnostic model. The masked set was used to test the effectiveness of the tree by calculating sensitivity and specificity.Student's t-test was used to compare peak intensities, because the distributions of values for protein peaks were Gaussian. Biomarker Pattern Software (Ciphergen Biosystems) was used to make a classification and decision tree (CART) (10Breiman L. Friedman J.H. Olshen R.A. Stone C.J. Classification and regression trees. Wadsworth, Belmont, CA1984Google Scholar), using those peaks with different expressions. In the tree, the spectrum pattern is classified according to a sequence of attributes which constitute nodes successively linked to one another. Each node has an expanding attribute to split the node into a number of branches. The classification of a specific profiling begins from the root node and extends to the terminal node, based on the answer to the question in the node. At each node of the tree, the question makes the data in the descendant node as "pure" as possible. For this purpose, the Gini index (11Rastogi R. Shim K. PUBLIC: a decision tree classifier that integrates building and pruning. 1998: 404-415Google Scholar) was used and defined as:Gini Index: Σk=12p^mk(1−p^mk) where p^mk is the fraction of controls in the node. The rationale for selecting the protein peak and the cut value was to minimize the sum of the Gini index of the whole nodes.To test the reproducibility of spectra, we ran one plasma sample in the same chip and in a different chip. Both the coefficient of variation (CV) for intensity and mass/charge (m/z) were calculated, based on duplicate sample testing. The intrachip CV for intensity was 1.8%, and the interchip CV was 2.1%. Both the intrachip and interchip CV for m/z were <0.05%. These values showed good reproducibility of spectra.Eighty-six protein peaks were identified in each plasma sample. The spectra between the endometriosis and control groups in the training set were quite different. The difference between the endometriosis and control groups was significant for 20 protein peaks (P<.05). These 20 protein peaks were used to construct a classification and decision tree. The four-layer tree was composed of three protein peaks with m/z values of 3,956.83, 11,710.70, and 6,986.45, to generate four terminal nodes (Fig. 1). For the training set, the tree showed a high sensitivity (32/36; 88.9%) and specificity (28/32; 87.5%) in the detection of endometriosis, while in the masked set, the tree as a diagnostic model showed a similar sensitivity of 87.5%, and a specificity of 85.7%.The diagnosis of endometriosis is still a challenge for any gynecologist. Several studies used serum markers such as CA125 (12Abrao M.S. Podgaec S. Filho B.M. Ramos L.O. Pinotti J.A. de Oliveira R.M. The use of biochemical markers in the diagnosis of pelvic endometriosis.Hum Reprod. 1997; 12: 2523-2527Crossref PubMed Scopus (87) Google Scholar, 13Guerriero S. Mais V. Ajossa S. Paoletti A.M. Angiolucci M. Melis G.B. Transvaginal ultrasonography combined with CA125 plasma levels in the diagnosis of endometrioma.Fertil Steril. 1996; 65: 293-298PubMed Scopus (74) Google Scholar), placental protein 14 (14Spuijbroek M.D. Dunselman G.A. Menheere P.P. Evers J.L. Early endometriosis invades the extracellular matrix.Fertil Steril. 1992; 58: 929-933Abstract Full Text PDF PubMed Scopus (159) Google Scholar), carbonic anhydrase antibodies (15Pittway D.E. Serum markers of endometrium and endometriosis.in: Diamond M.P. Osteen K.G. Endometrium and endometriosis. Blackwell Science, London1997: 31-41Google Scholar), and interleukin-6 (16Bedaiwy M.A. Falcone T. Sharma R.K. Goldberg J.M. Attaran M. Nelson D.R. et al.Prediction of endometriosis with serum and peritoneal fluid markers: a prospective controlled trial.Hum Reprod. 2002; 17: 426-431Crossref PubMed Scopus (381) Google Scholar) for the diagnosis of endometriosis. However, none of these showed both high sensitivity and high specificity in the detection of endometriosis. surface-enhanced laser desorption and ionization time-of-flight mass spectrometry uses patented protein chips to capture proteins with special characteristics in complex mixtures, and then analyze them by time-of-flight mass spectrometry to generate protein profiling. This novel technique can detect proteins with very low molecular weights (2,000–20,000 Da), and generate biomarker patterns composed of multiple protein markers.With the use of SELDI-TOF MS, we detected 20 protein peaks that were up- or down-regulated in the plasma of patients with endometriosis. We set up a tree for the classification of patients with endometriosis and healthy controls. The tree had a high sensitivity of 87.5%, and a specificity of 85.7%, in the unknown cases of the masked set. The results indicate that proteomic patterns detected by the SELDI system, coupled with biomarker pattern software, increase the sensitivity and specificity of the diagnosis of endometriosis. This new proteomic technique may be a useful tool in high-risk-population screening for endometriosis. Endometriosis is a common gynecological disease characterized by the growth of endometrial tissue outside the uterus, causing infertility and pelvic pain. The diagnosis of endometriosis is based on the histological identification of endometrial glands and stroma through invasive procedures such as laparoscopic surgery (1The American Fertility SocietyRevised American Fertility Society classification of endometriosis: 1985.Fertil Steril. 1985; 43: 351-352Abstract Full Text PDF PubMed Google Scholar). Up to now, there has been no reliable noninvasive method for detection of the disease, even though serum CA125 level has been widely used as a serological marker of endometriosis. In fact, the serum CA125 level is more useful in excluding epithelial cancer following the disease and in detecting recurrence (2Bast Jr, R.C. Xu F.J. Yu Y.H. Barnhill S. Zhang Z. Mill G.B. CA125: the past and the future.Int J Biol Markers. 1998; 13: 179-187Crossref PubMed Scopus (366) Google Scholar, 3Chen F.P. Soong Y.K. Lee N. Lo S.K. The use of serum CA125 as a marker for endometriosis in patients with dysmenorrhea for monitoring therapy and for recurrence of endometriosis.Acta Obstet Gynecol Scand. 1998; 77: 665-670Crossref PubMed Scopus (55) Google Scholar) than as a diagnostic method. A noninvasive and simple diagnostic method is urgently needed in both clinical practice and basic research. Recently developed proteomic techniques, such as surface-enhanced laser desorption and ionization time-of-flight mass spectrometry (SELDI-TOF MS), have been introduced for biomarker discovery. The SELDI-TOF MS system consists of a protein chip, mass analyzer, and bioinformatics software (4Chapman K. The Protein Chip Biomarker System from Ciphergen Biosystems: a novel proteomics platform for rapid biomarker discovery and validation.Biochem Soc Trans. 2002; 30: 82-87Crossref PubMed Google Scholar). The protein chips contain spots with different types of chromatographic material, i.e., hydrophobic, hydrophilic, ion-exchange, and immobilized metallic ion matrices. Small amounts of protein combine directly with the biochip by affinity capture, and are then analyzed by TOF MS to generate protein-profiling spectra. Surface-enhanced laser desorption and ionization enable a direct analysis of human body fluid with high speed, high throughput, and easy control. The new approach was successfully used in distinguishing the serum of healthy individuals from the serum of those with malignant diseases of the ovary (5Petricoin E.F. Ardekani A.M. Hitt B.A. Levine P.J. Fusaro V.A. Steinberg S.M. et al.Use of proteomic patterns in serum to identify ovarian cancer.Lancet. 2002; 359: 572-577Abstract Full Text Full Text PDF PubMed Scopus (2837) Google Scholar), prostate gland (6Qu Y.S. Adam B.L. Yutaka Y. Ward M.D. Cazares L.H. Schellhammer P.F. et al.Boosted decision tree analysis of surface-enhanced laser desorption/ionization mass spectral serum profiles discriminates prostate cancer from noncancer patients.Clin Chem. 2002; 48: 1835-1843PubMed Google Scholar), bladder (7Vlahou A. Schellhammer P.F. Mendrinos S. Patel K. Kondylis F.I. Gong L. et al.Development of a novel proteomic approach for the detection of transitional cell carcinoma of the bladder in urine.Am J Pathol. 2001; 158: 1491-1501Abstract Full Text Full Text PDF PubMed Scopus (393) Google Scholar), liver (8Poon T.C. Yip T.T. Chan A.T. Yip C. Yip V. Mok T.S. et al.Comprehensive proteomic profiling identifies serum proteomic signatures for detection of hepatocellular carcinoma and its subtypes.Clin Chem. 2003; 49: 752-760Crossref PubMed Scopus (233) Google Scholar), and lung (9Xiao X. Liu D. Tang Y. Guo F. Xia L. Liu J. et al.Development of proteomic patterns for detection lung cancer.Dis Markers. 2003; 19: 33-39Crossref PubMed Scopus (89) Google Scholar). The aim of this study was to determine the utility of plasma proteomic profiling with the use of SELDI-TOF MS for the detection of endometriosis. Eighty-seven patients who had been clinically diagnosed with endometriosis before surgery were enrolled in the study. All participants gave informed consent. All plasma samples were preoperatively collected. After surgery, 52 patients were histologically diagnosed with endometriosis, and their plasma samples were used for SELDI-TOF MS analysis. These patients were of reproductive age (range, 24–46 years), and had normal menstrual cycles. The staging of endometriosis was determined according to the revised scoring system of the American Fertility Society (1The American Fertility SocietyRevised American Fertility Society classification of endometriosis: 1985.Fertil Steril. 1985; 43: 351-352Abstract Full Text PDF PubMed Google Scholar). Plasma samples were taken from 46 female, age-matched blood donors from the blood transfusion department of Peking Union Medical College Hospital, Beijing, China. The blood donors were normal according to pelvic examination, vaginal sonography, and serum CA125 level. This study was approved by the Ethics Committee of Peking Union Medical College. All samples were obtained by venipuncture, and drawn into vacutainers with 3.2% sodium citrate and centrifuged at 3,000 rpm for 8 minutes at room temperature. The plasma was aliquoted and stored at −80°C until use. Plasma samples were thawed on ice and centrifuged at 10,000 rpm for 10 minutes. Twenty microliters of U9 buffer (9 M urea, 2% 3-[3-(Cholamidopropyl)dimethylammonio]-1-proanesulfonate (CHAPS), and 50 mM Tris-HCl, pH 9) were added to 10 μL of each plasma sample, and the solutions were vortexed for 30 minutes. The resulting plasma solutions were further diluted by 360 μL of buffer solution (50 mM potassium acetate, pH 4.0). Protein chips (WAX2, Ciphergen Biosystems, Freemont, CA) were preequilibrated with 200 μL of buffer solution twice. One hundred microliters of diluted plasma sample were added to each spot of WAX2 protein chip in a bioprocessor (Ciphergen Biosystems). The bioprocessor was then sealed and agitated on a platform shaker for 60 minutes. After discarding the remaining samples, the chips were washed with 200 μL of buffer solution for 5 minutes. This procedure was repeated three times, with two brief water rinses between applications of buffer solution. The chips were removed from the bioprocessor and air-dried. Each spot on the chip received two applications of 0.5 μL of a saturated solution of α-cyano-4-hydroxycinnamic acid in 50% v/v acetontrile and 5% v/v trifluoroacetic acid, and the chips were air-dried. All chips were analyzed with the use of the protein biological system II (PBS II) Protein Chip Reader (Ciphergen Biosystems). Data were collected by an average of 150 laser shots, with a detector sensitivity of 8 and a laser intensity of 195. The highest mass for detection was 50,000 Daltons (Da), with an optimization range between 2,000–20,000 Da. Mass accuracy was calibrated using the All-in-One Peptide Molecular Mass Standard (Ciphergen Biosystems). The SELDI-TOF mass spectra were obtained and analyzed using Ciphergen Protein Chip Software 3.1.1 (Ciphergen Biosystems). All spectra were compiled, and peak intensities were normalized in a mass range between 2,000–20,000 Da. The noise was filtrated using signal-to-noise ratio. An automatic peak detection program was used. Thirty-six samples from the endometriosis (EM) group and 32 samples from the control group were randomly selected as the training set. The remaining 16 EM samples and 14 control samples were designated as the masked set. The training set was used to compare the peak intensities of protein profiling between the two groups, and to set up a classification and regression tree as a diagnostic model. The masked set was used to test the effectiveness of the tree by calculating sensitivity and specificity. Student's t-test was used to compare peak intensities, because the distributions of values for protein peaks were Gaussian. Biomarker Pattern Software (Ciphergen Biosystems) was used to make a classification and decision tree (CART) (10Breiman L. Friedman J.H. Olshen R.A. Stone C.J. Classification and regression trees. Wadsworth, Belmont, CA1984Google Scholar), using those peaks with different expressions. In the tree, the spectrum pattern is classified according to a sequence of attributes which constitute nodes successively linked to one another. Each node has an expanding attribute to split the node into a number of branches. The classification of a specific profiling begins from the root node and extends to the terminal node, based on the answer to the question in the node. At each node of the tree, the question makes the data in the descendant node as "pure" as possible. For this purpose, the Gini index (11Rastogi R. Shim K. PUBLIC: a decision tree classifier that integrates building and pruning. 1998: 404-415Google Scholar) was used and defined as:Gini Index: Σk=12p^mk(1−p^mk) where p^mk is the fraction of controls in the node. The rationale for selecting the protein peak and the cut value was to minimize the sum of the Gini index of the whole nodes. To test the reproducibility of spectra, we ran one plasma sample in the same chip and in a different chip. Both the coefficient of variation (CV) for intensity and mass/charge (m/z) were calculated, based on duplicate sample testing. The intrachip CV for intensity was 1.8%, and the interchip CV was 2.1%. Both the intrachip and interchip CV for m/z were <0.05%. These values showed good reproducibility of spectra. Eighty-six protein peaks were identified in each plasma sample. The spectra between the endometriosis and control groups in the training set were quite different. The difference between the endometriosis and control groups was significant for 20 protein peaks (P<.05). These 20 protein peaks were used to construct a classification and decision tree. The four-layer tree was composed of three protein peaks with m/z values of 3,956.83, 11,710.70, and 6,986.45, to generate four terminal nodes (Fig. 1). For the training set, the tree showed a high sensitivity (32/36; 88.9%) and specificity (28/32; 87.5%) in the detection of endometriosis, while in the masked set, the tree as a diagnostic model showed a similar sensitivity of 87.5%, and a specificity of 85.7%. The diagnosis of endometriosis is still a challenge for any gynecologist. Several studies used serum markers such as CA125 (12Abrao M.S. Podgaec S. Filho B.M. Ramos L.O. Pinotti J.A. de Oliveira R.M. The use of biochemical markers in the diagnosis of pelvic endometriosis.Hum Reprod. 1997; 12: 2523-2527Crossref PubMed Scopus (87) Google Scholar, 13Guerriero S. Mais V. Ajossa S. Paoletti A.M. Angiolucci M. Melis G.B. Transvaginal ultrasonography combined with CA125 plasma levels in the diagnosis of endometrioma.Fertil Steril. 1996; 65: 293-298PubMed Scopus (74) Google Scholar), placental protein 14 (14Spuijbroek M.D. Dunselman G.A. Menheere P.P. Evers J.L. Early endometriosis invades the extracellular matrix.Fertil Steril. 1992; 58: 929-933Abstract Full Text PDF PubMed Scopus (159) Google Scholar), carbonic anhydrase antibodies (15Pittway D.E. Serum markers of endometrium and endometriosis.in: Diamond M.P. Osteen K.G. Endometrium and endometriosis. Blackwell Science, London1997: 31-41Google Scholar), and interleukin-6 (16Bedaiwy M.A. Falcone T. Sharma R.K. Goldberg J.M. Attaran M. Nelson D.R. et al.Prediction of endometriosis with serum and peritoneal fluid markers: a prospective controlled trial.Hum Reprod. 2002; 17: 426-431Crossref PubMed Scopus (381) Google Scholar) for the diagnosis of endometriosis. However, none of these showed both high sensitivity and high specificity in the detection of endometriosis. surface-enhanced laser desorption and ionization time-of-flight mass spectrometry uses patented protein chips to capture proteins with special characteristics in complex mixtures, and then analyze them by time-of-flight mass spectrometry to generate protein profiling. This novel technique can detect proteins with very low molecular weights (2,000–20,000 Da), and generate biomarker patterns composed of multiple protein markers. With the use of SELDI-TOF MS, we detected 20 protein peaks that were up- or down-regulated in the plasma of patients with endometriosis. We set up a tree for the classification of patients with endometriosis and healthy controls. The tree had a high sensitivity of 87.5%, and a specificity of 85.7%, in the unknown cases of the masked set. The results indicate that proteomic patterns detected by the SELDI system, coupled with biomarker pattern software, increase the sensitivity and specificity of the diagnosis of endometriosis. This new proteomic technique may be a useful tool in high-risk-population screening for endometriosis.