Background/Objectives: Kidney cancer (RC) is a significant global health burden. Renal cell carcinoma (RCC) is the most common form of kidney cancer. Its predominant histological subtype is clear cell renal cell carcinoma (ccRCC), which is frequently diagnosed at an advanced local stage or with metastases. Detecting cancer at an early stage significantly increases the likelihood of a cure; therefore, research on new markers and a thorough understanding of tumor biology are essential. This study investigated the significance of aromatase (ARO), cathepsin S (CTSS), and matrix metalloproteinase 1 (MMP-1) as potential biomarkers in ccRCC. Methods: ARO, CTSS, and MMP-1 concentrations in plasma were determined using SPRi biosensors. Appropriate antibodies were used as biorecognition molecules in the biosensors. The samples analyzed came from 60 patients with histopathologically confirmed clear cell renal cell carcinoma (ccRCC) and from 26 patients diagnosed with chronic cystitis or benign prostatic hyperplasia (BPH). Results: A statistically significant increase (p < 0.00001) in the concentration of all proteins compared with the control samples was observed at the T3-T4 stage. The ARO concentration was already statistically significantly higher at the T1-T2 stage (p < 0.00001). The ROC curve for aromatase demonstrated high sensitivity and specificity for detecting ccRCC, with a cut-off point of 7.53 ng mL-1. A moderate positive correlation was also found between the concentrations of the three tested substances in renal cancer, which may indicate potential interactions in the tumor's pathogenesis. Conclusions: SPRI testing has been shown to be an alternative to standard methods for detecting potential ccRCC markers. The biosensors used in the study can simultaneously determine ARO, CTSS, and MMP-1. The results obtained suggest the potential importance of these proteins in the development of ccRCC, and our work proposes a new diagnostic technique that may aid in the diagnosis of ccRCC.

Osteopontin (OPN) is a protein that plays many essential functions in the human body. It is present in most tissues and body fluids. OPN, among other things, participates in wound healing, the formation and remodeling of bone, immune response, inflammation, angiogenesis, and tumor formation. A new analytical method, based on SPRi (surface plasmon resonance imaging) biosensors, has been developed to determine osteopontin in biological fluids. OPN was captured from a solution by an immobilized antibody (mouse or rabbit), a bioreceptor in the SPRi sensor. A separate validation process was carried out for each antibody used. The LOD and LOQ values obtained for the biosensor with mouse antibody were 0.014 ng mL-1 and 0.043 ng mL-1, respectively, and those obtained for the biosensor with rabbit antibody were 0.018 ng mL-1 and 0.055 ng mL-1, respectively. The response ranges of both biosensors were in a similar range: 0.05-1.00 ng mL-1. OPN was determined in blood plasma to demonstrate the sensor potential, showing good agreement with the data obtained using an ELISA test and reported in the literature. The presented method is characterized by ease and speed of measurement, and the process does not require special preparation of samples.

Purpose Endometriosis is a prevalent gynecological condition often requiring invasive procedures for diagnosis. This study aimed to assess mesothelin (MSLN) as a potential non-invasive biomarker by analyzing its levels in peritoneal fluid (PF) and plasma. Materials/methods In a multicenter cross-sectional study, MSLN concentrations were measured using Surface Plasmon Resonance imaging (SPRi) biosensors in samples from women undergoing laparoscopy for pelvic pain or infertility. Results Significantly higher MSLN levels were observed in the PF of patients with endometriosis compared to controls (mean concentrations: 10.24 vs. 9.39 ng/mL, p = 0.038). The elevation was particularly notable in patients with stage I and II endometriosis compared to the control group (mean concentrations: 10.83 vs. 9.39 ng/mL, p = 0.049). No significant differences were observed in plasma. Conclusions Mesothelin in PF may be considered as a potential biomarker for endometriosis, especially in its early stages. Further studies are needed to confirm its diagnostic value.

Indoleamine 2,3-dioxygenase 1 (IDO-1) and interferon-gamma (IFN-γ) are proteins that play a significant role in inflammatory conditions and tumor development. The detection of IDO1 and IFN-γ is crucial for understanding their interplay in immune responses. This study introduced a novel method for the simultaneous quantitative determination of IDO-1 and IFN-γ in different biological samples/materials. The method is based on an optical biosensor, with surface plasmon resonance detection carried out by the imaging version of the sensor (SPRi). Biotinylated antibodies immobilized on the surfaces of the linker and carboxymethylated dextran served as the recognition elements for the developed biosensor. Relevant studies were conducted to optimize the activities of the biosensor by employing appropriate reagent concentrations. Validation was performed for each protein separately; low detection and quantification limits were obtained (for IDO-1 LOD = 0.27 ng/mL, LOQ = 0.81 ng/mL; for IFN-γ LOD = 1.76 pg/mL and LOQ = 5.29 pg/mL). The sensor operating ranges were 0.001–10 ng/mL for IDO-1 and 0.1–1000 pg/mL for IFN-γ. The constructed biosensor demonstrated its sensitivity and precision when the appropriate analytical parameters were determined, based on the proposed method. It can also selectively capture IDO-1 and IFN-γ from a large sample matrix. The biosensor efficiency was confirmed by the determination of IDO-1 and IFN-γ in simultaneous measurements of the plasma and urine samples of patients diagnosed with bladder cancer and the control group. The outcomes were compared to those obtained using a certified ELISA test, demonstrating convergence between the two methodologies. The preliminary findings demonstrate the biosensor’s efficacy and suitability for comprehensive analyses of the examined biological samples.

This study aimed to evaluate the concentration of osteopontin in peritoneal fluid and plasma as potential biomarkers for diagnosing endometriosis. Osteopontin levels were measured using surface plasmon resonance imaging (SPRI) biosensors in patients suspected of having endometriosis. Plasma samples were collected from 120 patients, and peritoneal fluid was collected from 86 patients. Based on the detection of endometriosis lesions during laparoscopy, participants were divided into a study group (patients with endometriosis) and a control group (patients without endometriosis). The results showed no significant differences in plasma osteopontin levels between women with endometriosis and the control group (19.86 ± 6.72 ng/mL vs. 18.39 ± 4.46 ng/mL, p = 0.15). Similarly, peritoneal fluid osteopontin concentrations did not differ significantly between patients with and without endometriosis (19.04 ± 5.37 ng/mL vs. 17.87 ± 5.13 ng/mL, p = 0.29). Furthermore, osteopontin levels in both plasma and peritoneal fluid were not significantly associated with the stage of endometriosis, the presence of endometrioma, or the menstrual cycle phase. The findings of this study do not support osteopontin concentration as a reliable biomarker for endometriosis. However, further research is necessary to explore osteopontin’s potential role in the disease.

Laminin, fibronectin, and collagen IV are pivotal extracellular matrix (ECM) components. The ECM environment governs the fundamental properties of tumors, including proliferation, vascularization, and invasion. Given the critical role of cell-matrix adhesion in malignant tumor progression, we hypothesize that the concentrations of these proteins may be altered in the plasma of patients with clear cell renal cell carcinoma (ccRCC). This study aimed to evaluate the serum, urine, and tissue levels of laminin-5, collagen IV, and fibronectin among a control group and ccRCC patients, with the latter divided into stages T1–T2 and T3–T4 according to the TNM classification. We included 60 patients with histopathologically confirmed ccRCC and 26 patients diagnosed with chronic cystitis or benign prostatic hyperplasia (BPH). Collagen IV, laminin-5, and fibronectin were detected using Surface Plasmon Resonance Imaging biosensors. Significant differences were observed between the control group and ccRCC patients, as well as between the T1–T2 and T3–T4 subgroups. Levels were generally higher in plasma and tissue for fibronectin and collagen IV in ccRCC patients and lower for laminin. The ROC (Receiver operating characteristic) analysis yielded satisfactory results for differentiating between ccRCC patients and controls (AUC 0.84–0.93), with statistical significance for both fibronectin and laminin in plasma and urine. Analysis between the T1–T2 and T3–T4 groups revealed interesting findings for all examined substances in plasma (AUC 0.8–0.95). The results suggest a positive correlation between fibronectin and collagen levels and ccRCC staging, while laminin shows a negative correlation, implying a potential protective role. The relationship between plasma and urine concentrations of these biomarkers may be instrumental for tumor detection and staging, thereby streamlining therapeutic decision-making.

BackgroundThe aim of this study was to analyze the concentration of leptin in peritoneal fluid and plasma and to assess their role as potential biomarkers in the diagnosis of endometriosis.Materials & methodsLeptin adjusted for BMI (leptin/BMI ratio) was measured using surface plasmon resonance imaging (SPRI) biosensors. Patients with suspected endometriosis were included in the study. Plasma was collected from 70 cases, and peritoneal fluid from 67 cases. Based on the presence of endometriosis lesions detected during laparoscopy, patients were divided into a study group and a control group (patients without endometriosis).ResultsLeptin/BMI ratio in plasma did not differ between women with endometriosis and the control group (0.7159 ± 0.259 vs 0.6992 ± 0.273, p= 0,7988). No significant differences were observed in peritoneal leptin/BMI ratio levels in patients with and without endometriosis (0.6206 ± 0.258 vs 0.6215 ± 0.264, p= 0,9896). Plasma and peritoneal leptin/BMI ratios were significantly lower in women with endometriosis - related primary infertility compared to women with endometriosis without primary infertility (0.640 ± 0.502 vs 0.878 ± 0.623, p < 0.05). The difference was observed in case of primary infertility, but not in terms of the secondary one. No significant differences were noted between leptin/BMI ratio in the proliferative phase and the secretory phase (0.716 ± 0.252 vs 0.697 ± 0.288, p= 0,7785).ConclusionThe results of present study do not support the relevance of leptin concentration determination as a biomarker of the endometriosis. Due to the limited number of samples in the tested group, further studies are needed to confirm its role.

The urothelial basement membrane (UBM) contains type IV collagen, laminin-5, and fibronectin. Urothelial neoplasm can break through the UBM underlying the urothelium to invade the lamina propria. Conceptually, all bladder cancer staging over Ta (T1-T4) may demonstrate disturbances in the UBM structure, as well as alterations in the serum concentrations of the studied components. The aim of this study was to determine the blood serum concentration of collagen IV, laminin-5 and fibronectin in bladder cancer patients. Quantification of their concentrations and correlation with various clinicopathological parameters may be useful for making more accurate predictions and identifying high-risk patients. The study included 96 patients with bladder cancer confirmed by transurethral resection or cystectomy and 26 patients with diagnosed cystitis chronica or BPH (benign prostate hyperplasia). Collagen IV, laminin-5 and fibronectin were detected using Surface Plasmon Resonance Imaging biosensors. Significant differences in blood serum concentrations of the studied biomarkers were observed between the control group and bladder cancer patients, as well as between nonmuscle-invasive and muscle-invasive groups. ROC analysis gave satisfactory results for differentiation between the control group and bladder cancer group (AUC 0.92-0.99), with lower values only for collagen IV between nonmuscle-invasive and muscle-invasive patients (AUC 0.71), and a statistically insignificant difference for laminin-5. Laminin-5 concentration was more closely correlated to tumour grade, size and recurrence rate; fibronectin to tumour stage, size and morphology; and collagen IV to tumour stage, grade and recurrence rate. The relations between serum concentrations of the presented biomarkers of the urothelial basement membrane may be useful for bladder cancer detection and for determination of the tumour stage, hence simplifying the making of therapeutic decisions.

Neuropilin-1 is transmembrane protein with soluble isoforms. It plays a pivotal role in both physiological and pathological processes. NRP-1 is involved in the immune response, formation of neuronal circuits, angiogenesis, survival and migration of cells. The specific SPRI biosensor for the determination of neuropilin-1 was constructed using mouse monoclonal antibody that captures unbound NRP-1 form body fluids. The biosensor exhibits linearity of the analytical signal between 0.01 and 2.5 ng/mL, average precision value 4.7% and recovery between 97% and 104%. The detection limit is 0.011 ng/mL, and the limit of quantification is 0.038 ng/mL. The biosensor was validated by parallel determination of NRP-1 in serum and saliva samples using the ELISA test, with good agreement of the results.

Follicle-stimulating hormone (FSH) regulates the development, growth, pubertal maturation and reproductive processes of the human body. The determination of serous FSH concentration is significant as an alternative to testicular biopsy in the case of boys suffering from cryptorchidism after orchidopexy, and as a means of determining the menopausal stage in women. The aim of this investigation is to develop a specific array surface plasmon resonance imaging (SPRi) biosensor for the determination of FSH in body liquids such as blood plasma, obtaining sufficient sensitivity to determine FSH at levels characteristic for that hormone in blood plasma, without any signal enhancement. The biosensor consists of a mouse monoclonal anti-FSH antibody attached to the gold surface of a chip via a cysteamine linker. Its linear response range is from 0.08 mIU mL-1 (LOQ) to 20 mIU mL-1, and well covers most of the range of FSH activities found in blood without dilution. The precision of measurement is between 3.2% and 13.1% for model samples, and between 3.7% and 5.6% for spiked plasma samples. Recoveries are in the range from 94% to 108%. The biosensor has good selectivity, and is validated by comparison with ECLE, with good agreement of the results

Research on the markers of immunoregulatory response in multiple sclerosis (MS) is still of great importance. The aim of our study was the evaluation of leptin, fibronectin, and UCHL1 concentrations as potential biomarkers of a relapsing-remitting type of MS (RRMS). Surface Plasmon Resonance Imaging (SPRI) biosensors were used for the evaluation of proteins concentrations in 100 RRMS patients and 46 healthy volunteers. Plasma leptin, fibronectin, and UCHL1 concentrations were significantly higher in RRMS patients compared to the control group (p < 0.001, respectively). UCHL1 concentration evaluation revealed the highest diagnostic sensitivity (100%) and negative predictive value (100%) in differentiating MS patients from healthy individuals. There was no significant difference in the UCHL1 concentrations depending on the patient's sex, the presence of relapse within the last 24 months, and the EDSS value (p > 0.05, respectively). In RRMS patients UCHL1 concentration positively correlated with fibronectin levels (r = 0.3928; p < 0.001). In the current cohort of patients plasma UCHL1 concentration was independent of the time of MS relapse and the severity of neurological symptoms. Thus current study may indicate that plasma UCHL1, besides leptin and fibronectin, also could be a promising high-sensitive potential biomarker of relapsing-remitting type of MS. However, these results should be validated with a larger group of patients, taking into account neuroimaging and cerebrospinal fluid analysis data, and by comparing them to patients with other neurological diseases as a control group.

Cortisol is a hormone which plays an essential role in the immune, endocrine, cardiovascular, renal and skeletal systems. Its level increases in response to stress, illness, injury or exhaustion, and it is therefore a significant diagnostic biomarker of stress. An immunosensor for the determination of cortisol by SPRi array was developed. The receptive part of the immunosensor is mouse monoclonal antibody against cortisol, immobilized via cysteamine linker. The optimum pH of the immunosensor is 7.4, and the optimum concentration of the antibody is 50 ng mL−1. The immunosensor is specific for cortisol, and its linear response ranges from 0.20 ng mL−1 (LOQ) to 8 ng mL−1. The precision of the determination was between 3.1% and 3.3%, and the recovery between 99% and 102%. The immunosensor was validated by simultaneous determination of cortisol in serum and saliva samples by a standard method, with good agreement between the results.

The ubiquitin–proteasome system (UPS) participates in the degradation of proteins which play an important role in regulating the cell cycle, apoptosis, and angiogenesis, as well as in the immune system. These processes are important in carcinogenesis. Transitional cell carcinoma (TCC) is one of the predominant types of bladder cancer. The relationship between the ubiquitin–proteasome system and cancer progression has become a topic of increasing interest among researchers. In this work, we propose an application of surface plasmon resonance imaging (SPRi)-based biosensors for the detection of 20S proteasome and ubiquitin carboxy-terminal hydrolase L1 (UCH-L1) in the blood serum and urine of patients with TCC. The aim of the study was to determine 20S proteasome and UCH-L1 concentrations and to correlate the results with clinicopathological parameters. The group of subjects consisted of 82 patients with confirmed TCC, in addition to a control group of 27 healthy volunteers. It was found that 20S proteasome and UCH-L1 concentrations were significantly elevated in both the serum and urine of TCC patients, compared with the healthy subjects. There was a correlation between 20S proteasome concentrations in serum and urine, as well as between serum proteasome and UCH-L1 concentration. The SPRi biosensor sensitive to 20S proteasome using PSI inhibitor as the receptor, and the SPRi biosensor sensitive to the UCH-L1 protein using the protein-specific antibody as the receptor is suitable for the determination of 20S proteasome and UCH-L1 in body fluids and can serve as useful tools in the investigation of cancer biomarkers.

Cathepsin S is an emerging marker for ovarian cancer. Two 'analytically specific' SPRi biosensors for the determination of Cath S have been developed. The reception part of one of the biosensors consists of the rat monoclonal antibody specific for cathepsin S attached to the gold surface via covalent bonds with cysteamine linker, while the second biosensor consists of the inhibitor LY3000328 attached via hydrophobic interaction with the 1-octadecanothiol linker. Under optimized conditions, in terms of pH and receptor concentration, both biosensors have linear response ranges between LOQ (0.14 ng mL(-1)) and 2.5 ng mL(-1), which is suitable for the determination of Cath S in blood plasma samples of ovarian cancer patients and healthy individuals, after corresponding dilution with 0.15 M PBS buffer. Precision and recoveries are quite acceptable: below 7% and 98-101% respectively for the biosensor with antibody, and below 12% and 101-103% for the biosensor with inhibitor. The biosensors were validated by the determination of Cath S in series of plasma from ovarian cancer patients and healthy volunteers using both biosensors and ELISA, giving Pearson coefficients close to 1. Plasma Cath S concentration can be used as an ovarian cancer marker, in view of the highly elevated concentrations detected.

Leptin is a hormone that has a fundamental role in the regulation of feeding and energy balance. A developed specific SPRi immunosensor for leptin may be a new tool for leptin determination in blood plasma. The immunosensor consists of rabbit anti-leptin antibody immobilized on a gold chip via cysteamine linker, using the EDC/NHS protocol. Non-fluidic array SPRi is used for analytical signal formation. Under optimized conditions, the linear response range of the immunosensor covers concentrations from 0.23 to 5 ng mL(-1). The LOD of the immunosensor is 0.07 ng mL(-1), and the LOQ is 0.23 ng mL(-1). The precision of measurement depends on leptin concentration, and is between 9.1% and 2.2%. Recoveries of the leptin spike are between 97% and 110%. The immunosensor and related analytical method were validated by parallel determination of leptin in series of plasma from children suffering from malnutrition and a control group, using Enzyme-Linked Immunosorbent Assay (ELISA) and SPRi. Pearson's correlation coefficient was equal to 0.991. The developed immunosensor and related method are more direct, faster and much simpler than ELISA.

Objectives: Composition of saliva reflects the condition of the oral cavity. The aim of the study: Investigation of the concentrations of MMP-1 (Matrix metalloproteinase-1), MMP-2 (Matrix metalloproteinase-2) and fibronectin in the saliva of patients planned for endodontic treatment or surgical extraction. Material and methods: Seventy-five patients with caries and 14 healthy subjects were included in the study. Subjects were divided into group 1, in which 50 patients were planned for endodontic treatment, and group 2, in which 25 patients were planned for surgical extraction. For the measurements, we used a surface plasmon resonance imaging biosensor. Results: We found higher levels of MMP-1, MMP-2 and fibronectin in the saliva of patients planned for dental treatment than in healthy donors. We found lower concentrations of MMP-2 in subjects planned for surgical extraction, than in patients planned for endodontic treatment; however, there were no such differences in salivary concentrations of MMP-1 and fibronectin. There were no statistically significant differences in MMP-1 concentrations in the saliva before and after any type of dental treatment, but contrary to that, we found a statistically significant decrease in MMP-2 concentrations after endodontic treatment and after surgical extraction. We found a significant rise in the concentrations of fibronectin after surgical extraction but not after endodontic treatment. Conclusions: The concentrations of MMP-1 and MMP-2 in the saliva of our patients with caries were increased in comparison to healthy individuals, but after the treatment-so sanation of the oral cavity-we noted a decrease in matrix metalloproteinases (MMPs) levels. MMPs can be found in gingival crevicular fluid and saliva, carious dentin and plaque. According to our observations, the main source of MMPs in patients with caries is probably carious dentin. Increase in the salivary levels of fibronectin (FN) after surgical extraction may be connected with soft tissue injury caused by surgical extraction. Our results are another example of the fact that higher salivary concentrations of MMP-1, MMP-2 and FN can reflect the health status of the oral cavity in patients with caries.

Purpose: Laminin 5 and collagen IV are the main compounds of the extracellular matrix of the germinal epithelium. The purpose of this study was to evaluate the concentration of these two markers of fibrosis in the plasma of boys with congenital unilateral cryptorchidism. Materials and methods: The study group comprised 43 boys aged 1-3 years with congenital unilateral cryptorchidism. The control group included 54 healthy, age matched boys, admitted for planned hernioplasty. To assess laminin 5 and collagen IV in the plasma of boys with unilateral cryptorchidism, we used a new biosensor with Surface Plasmon Resonance Imaging technique detection. Results: The median concentration of laminin 5 and collagen IV in the serum of boys with congenital, unilateral cryptorchidism was higher than in boys with normal scrotal testis. The difference was statistically significant (p < 0.0001). We did not notice a correlation between a higher position of the testicles in the inguinal and/or their condition and levels of laminin 5 and collagen IV in the plasma. Conclusion: Laminin 5 and collagen IV concentrations in the plasma were higher in patients with congenital unilateral cryptorchidism. We believe that in the future, our results could be compared with fertility level in adulthood.

The 20S proteasome, released into the circulation, is a novel cancer biomarker. It exists in two forms: the constitutive proteasome (20Sc) and the immunoproteasome (20Si), which both have separate diagnostic significance. The aim of this work was to develop new methods for 20Si and 20Sc determination. Five alternative specific biosensors usable with the surface plasmon resonance imaging (SPRI) technique for 20Si determination have been developed. Specific 20Si entrapment on the biosensor surface from an analyzed solution was achieved by means of an immobilized specific 20Si receptor. Four of the biosensors contain newly synthesized specific 20Si receptors, while the fifth contains the inhibitor ONX 0914. A method for 20Sc determination using an SPRI biosensor containing PSI inhibitor has been developed. By the introduction of an inhibitor blocking 20Si, 20Sc is selectively determined. All of the methods developed for 20Si and 20Sc determination exhibit good selectivity and satisfactory precision, recoveries and dynamic response ranges. 20Si and 20Sc were determined in blood plasma samples from healthy donors and patients with acute leukemia. In the case of these patients 20Si was the major component, and its level was more than one order of magnitude higher than in the healthy donors.

The purpose of this study was to develop a new method for a determination of the cathepsin L-biosensor based on the Surface Plasmon Resonance Imaging technique. The cathepsin L is an endopeptidase, which degrades proteins and plays an important role in various processes occurring in the human body. The detection technique, Surface Plasmon Resonance Imaging, is an optical, label-free technique, which can be used for quantitative determination of the different proteins. In order to bind the enzyme, the cathepsin L inhibitor-RKLLW-NH2 was used. The validation process showed that parameters: precision, accuracy, and selectivity of the method were acceptable. The analytically useful range of the standard curve was 0.50 ng/mL-15.00 ng/mL. The detection and quantification limit of method was 1.67 pg/mL and 5.07 pg/mL, respectively. The usefulness of the developed method was confirmed by the determination of the cathepsin L concentration in the blood plasma of some healthy persons and in the blood plasma of patients. The obtained results were compared with the results obtained by the ELISA. It was found that the correlation between these two methods was very strong, what suggest that the developed method can be used as the competitive method to the ELISA.

The matrix metalloproteinases are enzymes capable of remodeling of extracellular matrix, and modulate the behavior of cells. Maturation of gubernaculum and spermatogenesis demand proper equilibrium of metalloproteinases and their inhibitors. The aim of this survey was to investigate the levels of matrix metalloproteinase type 1 (MMP-1) and matrix metalloproteinase type 2 (MMP-2) in the plasma of children with unilateral cryptorchidism along with levels of Insulin-like Peptide 3 (INSL3) and inhibin B. INSL3 have a role in gubernaculum development. Inhibin B is produced by Sertoli cells, and its levels reflect the status of the testis germinative epithelium. Fifty boys with an undescended testicle, aged 1-4 years (median = 2.4 years) were enrolled into the study. Fifty boys with inguinal hernia aged 1-4 years, served as controls (median age = 2.1 years). Investigators assessed the MMP-1 and MMP-2 concentrations using Surface Plasmon Resonance Imaging. The levels of INSL-3 and inhibin B were assessed using commercial enzyme-linked immunosorbent assay ELISA. The median concentration of MMP-1 and MMP-2 in the blood plasma of patients with unilateral cryptorchidism, was nearly 2-folds higher than in controls. The great area under the ROC curve with the cut off value of 0.865 for MMP-1, and 0.819 for MMP-2, indicates the high clinical sensitivity and specificity of the test of plasma levels of MMP-1 and MMP-2 for boys with cryptorchidism. The increased plasma levels of MMP-1 and MMP-2, probably reflect the level of apoptosis of the germ cells in undescended testicles, in response to the heat stress during the period of prepubertal testis development. In the group of cryptorchid boys, we found slightly lower concentrations of INSL3, without statistical significance and without correlation with MMP-1 and MMP-2 levels. There were no significant differences in the levels of inhibin B in the group of boys with cryptorchidism and boys with inguinal hernia and it also did not correlate with MMP-1 and MMP-2 concentrations.