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Research on the markers of autoimmune response in multiple sclerosis (MS) is still of great importance. The aim of our study was the evaluation of plasma 20S constitutive proteasome, 20S immunoproteasome, and cathepsin S concentrations as potential biomarkers of a relapsing-remitting type of MS (RRMS). Surface plasmon resonance imaging (SPRI) biosensors were used for the evaluation of protein concentrations. Plasma 20S constitutive proteasome, 20S immunoproteasome, and cathepsin S concentrations were significantly higher in RRMS patients compared to the control group. All three parameters were characterized by excellent usefulness in differentiating MS patients from healthy individuals (AUC equal to or close to 1.000). The plasma concentration of analyzed parameters was not correlated with severity of disability in the course of RRMS (EDSS value), the number of years from the first MS symptoms, the number of years from MS diagnosis, or the number of relapses within the 24-month observational period. Our study has shown that plasma concentrations of 20S constitutive proteasome, 20S immunoproteasome, and cathepsin S have promising potential in differentiating RRMS patients from healthy individuals. All of the analyzed parameters were found to be independent of the time of MS relapse and the severity of neurological symptoms. Hence, their potential as highly sensitive and independent circulating markers of RRMS suggests a stronger association with immunological activity (inflammatory processes) than with the severity of the disease.
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Background: The 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 & methods: Leptin 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). Results: Leptin/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). Conclusion: The 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.
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Endometriose , Infertilidade Feminina , Humanos , Feminino , Endometriose/patologia , Leptina , Índice de Massa Corporal , BiomarcadoresRESUMO
PURPOSE: Endometriosis is a common disease with a complex pathomechanism and atypical symptoms, often leading to delayed diagnosis. Currently, the sole method for confirming the presence of the disease is through laparoscopy and histopathological examination of collected tissue. However, this invasive procedure carries potential risk and complications, necessitating the exploration of non-surgical diagnostic methods for endometriosis. This study aims to analyze peritoneal fluid and plasma samples for the expression of cathepsin L and cathepsin S to identify potential biomarkers for non-invasive diagnostic approaches to endometriosis. MATERIAL AND METHODS: In this cross-sectional study, plasma and peritoneal fluid samples were obtained during laparoscopy from 63 patients diagnosed with chronic pelvic pain or infertility. The study group consisted of women with confirmed endometriosis. The concentrations of cathepsins L and S were determined using an SPRi biosensor. RESULTS: The study did not reveal significant differences in the concentrations of cathepsin L and cathepsin S between the control group and the study group, both in peritoneal fluid and plasma. CONCLUSIONS: Based on the results of this study, it appears that cathepsins L and S are not suitable candidates as biomarkers for endometriosis.
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Acute lymphoblastic leukemia (ALL) and glioma are some of the most common malignancies, with ALL most often affecting children and glioma affecting adult men. Proangiogenic cytokines and growth factors play an important role in the development of both of these tumors. Glioma is characterized by an extremely extensive network of blood vessels, which continues to expand mainly in the process of neoangiogenesis, the direct inducers of which are cytokines from the family of vascular endothelial growth factors, i.e., vascular endothelial growth factor (VEGF-A) and its receptor vascular endothelial growth factor receptor 2 (VEGF-R2), as well as a cytokine from the fibroblast growth factor family, fibroblast growth factor 2 (FGF-2 or bFGF). Growth factors are known primarily for their involvement in the progression and development of solid tumors, but there is evidence that local bone marrow angiogenesis and increased blood vessel density are also present in hematological malignancies, including leukemias. The aim of this study was to examine changes in the concentrations of VEGF-A, VEGF-R2, and FGF-2 (with a molecular weight of 17 kDa) in a group of patients divided into specific grades of malignancy (glioma) and a control group; changes of VEGF-A and FGF-2 concentrations in childhood acute lymphoblastic leukemia and a control group; and to determine correlations between the individual proteins as well as the influence of the patient's age, diet, and other conditions that may place the patient in the risk group. During the statistical analysis, significant differences in concentrations were found between the patient and control groups in samples from people with diagnosed glioma and from children with acute lymphoblastic leukemia, but in general, there are no significant differences in the concentrations of VEGF-A, VEGF-R2, and FGF-2 between different grades of glioma malignancy. Among individuals treated for glioma, there was no significant impact from the patient's gender and age, consumption of food from plastic packaging, frequency of eating vegetables and fruit, smoking of tobacco products, the intensity of physical exercise, or the general condition of the body (Karnofsky score) on the concentrations of the determined cytokines and receptor. The listed factors do not bring about an actual increase in the risk of developing brain glioma.
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Glioma , Leucemia-Linfoma Linfoblástico de Células Precursoras , Masculino , Adulto , Criança , Humanos , Fator A de Crescimento do Endotélio Vascular/metabolismo , Citocinas/metabolismo , Fator 2 de Crescimento de Fibroblastos/metabolismo , Fatores de Crescimento do Endotélio Vascular/metabolismo , Glioma/metabolismo , Encéfalo/metabolismoRESUMO
Alzheimer's disease is among the neurodegenerative diseases for which there is a lack of rapid, effective, and non-invasive diagnostic methods. The development of a phospho-Tau 181 assay biosensor is therefore a response to the need for methods to diagnose AD. The present work was aimed at developing a fast, selective, and repeatable method for the quantitative determination of phospho-Tau 181, which could be used even during routine blood tests. Our method is a form of what is called liquid biopsy. The developed method underwent validation, as a result of which its analytical parameters were determined. An LOQ of 3.35 pg mL-1 was obtained, confirming the possibility of trace analysis of phospho-Tau 181 in human plasma. Relative percentage error values below 15 % and CVs in the range 1.47-7.09 % attest to the high accuracy and precision of the presented method. Also, the sample matrix was not found to significantly affect the results obtained for phospho-Tau 181 concentrations. The new SPRi biosensor provides reproducible measurements of the analyte under study (CV = 3.18-4.26 %). Although the method requires absolute adherence to the recommendations of the analytical procedure protocol, it achieves high selectivity and provides 90 % certainty of the correctness of the diagnosis based on measurements of phospho-Tau 181 concentration.
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Doença de Alzheimer , Técnicas Biossensoriais , Humanos , Doença de Alzheimer/diagnóstico por imagem , Proteínas tau , Ressonância de Plasmônio de Superfície/métodos , Biomarcadores , Peptídeos beta-AmiloidesRESUMO
A new method for the determination of cadherin 12 (CDH12)-an adhesive protein that has a significant impact on the development, growth, and movement of cancer cells-was developed and validated. The method is based on a biosensor using surface plasmon resonance imaging (SPRi) detection. A quartz crystal microbalance was used to analyze the characteristics of the formation of successive layers of the biosensor, from the linker monolayer to the final capture of CDH12 from solution. The association equilibrium constant (KA = 1.66 × 1011 dm3 mol-1) and the dissociation equilibrium constant (KD = 7.52 × 10-12 mol dm-3) of the anti-CDH12 antibody-CDH12 protein complex were determined. The determined analytical parameters, namely the values determining the accuracy, precision, and repeatability of the method, do not exceed the permissible 20% deviations specified by the aforementioned institutions. The proposed method is also selective with respect to possible potential interferents, occurring in up to 100-fold excess concentration relative to the CDH12 concentration. The determined Limit of Quantification (LOQ = 4.92 pg mL-1) indicates the possibility of performing quantitative analysis in human plasma or peritoneal fluid without the need to concentrate the samples; however, particular attention should be paid to their storage conditions, as the analyte does not exhibit high stability. The Passing-Bablok regression model revealed good agreement between the reference method and the SPRi biosensor, with ρSpearman values of 0.961 and 0.925.
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Técnicas Biossensoriais , Ressonância de Plasmônio de Superfície , Humanos , Ressonância de Plasmônio de Superfície/métodos , Líquido Ascítico , Técnicas Biossensoriais/métodos , CaderinasRESUMO
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.
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Técnicas Biossensoriais , Líquidos Corporais , Masculino , Humanos , Feminino , Animais , Camundongos , Hormônio Foliculoestimulante , Ressonância de Plasmônio de Superfície/métodos , Técnicas Biossensoriais/métodos , PlasmaRESUMO
Fluidic and non-fluidic surface plasmon resonance measurements were realized for the same type of sensory layer and using the same mouse IgG antibody and anti-mouse IgG antibody biomolecular system. A comparison of the thicknesses of the anti-mouse IgG antibody layers bound to the ligand at increasing analyte concentrations ranging from 0.0 µg mL-1 to 5.0 µg mL-1 in the non-fluidic and the fluidic variant showed that the thickness of the bound anti-mouse antibody layers in the fluidic variant was approximately 1.5-3 times larger than in the non-fluidic variant. The greater thicknesses of the deposited layers were also reflected in the larger increment of the resonant angle in the fluidic variant compared to the non-fluidic variant in the considered range of analyte concentrations. The choice between fluidic and non-fluidic surface plasmon resonance biosensors may be justified by the availability of analyte volume and the intended modulation technique. When working with limited analyte, non-fluidic biosensors with intensity modulation are more advantageous. For larger analyte quantities, fluidic biosensors with angular modulation are recommended, primarily due to their slightly higher sensitivity in this measurement mode.
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Técnicas Biossensoriais , Ressonância de Plasmônio de Superfície , Animais , Camundongos , Ressonância de Plasmônio de Superfície/métodos , Técnicas Biossensoriais/métodos , Imunoglobulina G , LigantesRESUMO
Alzheimer's disease (AD) is the most common form of dementia in older people. Its prevalence is expected to increase, and therefore it poses a serious challenge to the healthcare system. The aim of the study was to assess the concentration of leptin, cystatin C, neuropilin-1 and tau protein, as well as the influence of dietary habits on these parameters, in a group of AD patients (n = 110) compared to 60 healthy people (n = 60). It has been shown that AD patients, compared to healthy people, are characterized by significantly higher median concentrations of leptin (9.97 vs. 3.08), cystatin c (1.53 vs. 0.56) and tau protein (8.46 vs. 4.19), but significantly lower median neuropilin-1 (69.94 vs. 167.28). Multiple regression analyses showed that leptin levels could be explained by dietary habits in 27%, cystatin C in 51%, neuropilin-1 in 41% and tau protein in 25% of cases. Modification of eating habits may contribute to improving the values of the discussed parameters.
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A new biosensor based on the "surface plasmon resonance imaging (SPRi)" detection technique for the quantification of "fibroblast growth factor 23 (FGF23)" has been developed. FGF23 is mainly produced in bone tissues as a phosphaturic hormone that forms a trimeric complex with "fibroblast growth factor receptor 1 (FGFR1)" and αKlotho upon secretion. FGF23 stimulates phosphate excretion and inhibits the formation of active vitamin D in the kidneys. FGF23 has been shown to play a role in bone carcinogenesis and metastasis. The newly developed method, based on the array SPRi biosensor, was validated-the precision, accuracy, and selectivity were acceptable, and yielded less than ±10% recovery. The rectilinear response of the biosensor ranges from 1 to 75 pg/mL. The limit of detection was 0.033 pg/mL, and the limit of quantification was 0.107 pg/mL. The biosensor was used to determine FGF23 concentrations in the blood plasma of healthy subjects and patients with "clear cell" renal cell carcinoma (ccRCC). The obtained results were compared with those measured through an "enzyme-linked immunosorbent assay (ELISA)". The determined Pearson correlation coefficients were 0.994 and 0.989, demonstrating that the newly developed biosensor can be used as a competitive method for the ELISA.
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Técnicas Biossensoriais , Ressonância de Plasmônio de Superfície , Humanos , Fator de Crescimento de Fibroblastos 23 , Técnicas Biossensoriais/métodos , Fosfatos/metabolismo , Ensaio de Imunoadsorção Enzimática , Fatores de Crescimento de Fibroblastos/metabolismoRESUMO
Endometriosis is a chronic disease in which the endometrium cells are located outside the uterine cavity. The aim of this study was to evaluate circulating 20S proteasome and 20S immunoproteasome levels in plasma and peritoneal fluid in women with and without endometriosis in order to assess their usefulness as biomarkers of disease. Concentrations were measured using surface plasmon resonance imaging biosensors. Patients with suspected endometriosis were included in the study-plasma was collected in 112 cases and peritoneal fluid in 75. Based on the presence of endometriosis lesions detected during laparoscopy, patients were divided into a study group (confirmed endometriosis) and a control group (patients without endometriosis). Proteasome and immunoproteasome levels in both the plasma (p = 0.174; p = 0.696, respectively) and the peritoneal fluid (p = 0.909; p = 0.284, respectively) did not differ between those groups. There was a statistically significant difference in the plasma proteasome levels between patients in the control group and those with mild (Stage I and II) endometriosis (p = 0.047) and in the plasma immunoproteasome levels in patients with ovarian cysts compared to those without (p = 0.017). The results of our study do not support the relevance of proteasome and immunoproteasome determination as biomarkers of the disease but suggest a potentially active role in the pathogenesis of endometriosis.
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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.
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Técnicas Biossensoriais , Ressonância de Plasmônio de Superfície , Animais , Camundongos , Ressonância de Plasmônio de Superfície/métodos , Neuropilina-1 , Técnicas Biossensoriais/métodos , Ensaio de Imunoadsorção Enzimática , Diagnóstico por ImagemRESUMO
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.
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Esclerose Múltipla , Humanos , Esclerose Múltipla/diagnóstico , Fibronectinas , Leptina , Nível de Saúde , Voluntários Saudáveis , Ubiquitina TiolesteraseRESUMO
A biosensor was developed for the quantification of poly(ADP-ribose) polymerase-1 (PARP-1) in body fluids. An antibody specific for PARP-1 was placed on a chip with cysteamine (linker) and a gold layer. This biosensor has a linear response range (10-1000 pgâmL-1) under appropriate pH conditions and with an antibody ligand concentration of 5 ngâmL-1. Plasma samples were diluted with PBS buffer in appropriate quantities so that they fell within the linear range of the calibration curve. The biosensor exhibited suitable precision and accuracy, and good recovery (at levels from 95% to 105%). The method was validated by means of PARP-1 determinations in plasma samples from patients with endometriosis and a control group, using surface plasmon resonance imaging (SPRi) biosensors and an enzyme-linked immunosorbent assay (ELISA) test. The Spearman correlation coefficient was close to 1. PARP-1 may be a marker providing information about pathological changes in the body during endometriosis.
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Diagnostics based on the determination of biomarkers in body fluids will be more successful when several biomarkers are determined. A multiple-array SPRi biosensor for the simultaneous determination of CA125, HE4, CEA, IL-6 and aromatase has been developed. Five individual biosensors were placed on the same chip. Each of them consisted of a suitable antibody covalently immobilized onto a gold chip surface via a cysteamine linker by means of the NHS/EDC protocol. The biosensor for IL-6 works in the pg mL-1 range, that for CA125 in the µg mL-1 range, and the other three within the ng mL-1 range; these are ranges suitable for the determination of biomarkers in real samples. The results obtained with the multiple-array biosensor are very similar to those obtained with a single biosensor. The applicability of the multiple biosensor was demonstrated using several examples of plasma from patients suffering from ovarian cancer and endometrial cyst. The average precision was 3.4% for the determination of CA125, 3.5% for HE4, 5.0% for CEA and IL-6, and 7.6% for aromatase. The simultaneous determination of several biomarkers may be an excellent tool for the screening of the population for earlier detection of diseases.
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Aromatase , Técnicas Biossensoriais , Feminino , Humanos , Biomarcadores Tumorais , Interleucina-6 , Proteína 2 do Domínio Central WAP de Quatro Dissulfetos , Técnicas Biossensoriais/métodosRESUMO
Circulating body fluids such as blood, urea, saliva, cerebrospinal fluid, etc [...].
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Técnicas Biossensoriais , Líquidos Corporais , Saliva , Biomarcadores , UreiaRESUMO
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.
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Técnicas Biossensoriais , Neoplasias da Bexiga Urinária , Masculino , Humanos , Neoplasias da Bexiga Urinária/cirurgia , Colágeno Tipo IV , Fibronectinas , Ressonância de Plasmônio de Superfície , Soro , Biomarcadores , CalininaRESUMO
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.
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Técnicas Biossensoriais , Animais , Camundongos , Técnicas Biossensoriais/métodos , Hidrocortisona , Imunoensaio/métodos , Saliva , AnticorposRESUMO
Laparoscopy as a diagnostic tool for patients with suspected endometriosis is associated with several potentially life-threatening complications. Therefore, it is imperative to identify reliable, non-invasive biomarkers of the disease. The aim of this study was to analyse the concentrations of fibronectin and type IV collagen in peritoneal fluid and plasma to assess their role as potential biomarkers in the diagnosis of endometriosis. Fibronectin and collagen IV protein levels were assessed by surface plasmon resonance imaging (SPRi) biosensors with the usage of monoclonal antibodies. All patients enrolled in the study were referred for laparoscopy for the diagnosis of infertility or chronic pelvic pain (n = 84). The study group included patients with endometriosis confirmed during surgery (n = 49). The concentration of fibronectin in the plasma (329.3 ± 98.5 mg/L) and peritoneal fluid (26.8 ± 11.1 µg/L) in women with endometriosis was significantly higher than in the control group (251.2 ± 84.0 mg/L, 7.0 ± 5.9 µg/L). Fibronectin levels were independent of endometriosis stage (p = 0.874, p = 0.469). No significant differences were observed in collagen IV levels (p = 0.385, p = 0.465). The presence of elevated levels of fibronectin may indicate abnormalities in cell-ECM signalling during the course of endometriosis, and may be a potential biomarker for early detection.
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Endometriose , Humanos , Feminino , Endometriose/metabolismo , Líquido Ascítico/metabolismo , Fibronectinas/metabolismo , Colágeno Tipo IV/metabolismo , Biomarcadores/metabolismoRESUMO
Brain gliomas constitute the vast majority of malignant tumors of the nervous system. There is still a lack of fast, reliable and non-invasive methods of diagnostics. Our work focuses on the quantification of cathepsin B, D and S in glioma. The research was conducted with the use of SPRi biosensors sensitive to individual cathepsins. Changes in the quantity of selected cathepsins (cathepsins B, D and S), depending on the advancement of glioma and the presence or absence of important features or comorbidities in the selected patient, were examined. The results were statistically analyzed and interpreted based on the available clinical description. Statistical significance was observed in the difference in the concentration of the studied cathepsins, mainly between the groups Control and G3/G4 and G1/G2 and G3/G4. The strength of the correlation between the concentrations of individual cathepsins and the age of the patient and the size of the tumor, as well as the correlation between individual proteins, was investigated. The influence of IDH 1/2 status on the concentration of determined cathepsins was investigated and ROC analysis was performed. As a result of our research, we have developed a method for the diagnosis of brain glioma that allows us to distinguish grades G1/G2 from G3/G4 and the control group from G3/G4. We found an average positive correlation between the concentrations of the proteins tested and the age of the patient and a high positive correlation between the cathepsins tested. Comparative analysis of the effect of the presence of IDH 1/2 mutations on the number of proteins tested allowed us to demonstrate that the cathepsins assayed can be independent markers.