The Cockcroft Gault formula is often used to calculate the glomerular filtration rate (GFR) from plasma creatinine results. In Sweden this calculation is not usually done in the laboratory, but locally in the wards. These manual calculations could cause erroneous results. In several studies plasma cystatin C has been shown to be superior to plasma creatinine for estimation of GFR. One limitation of using cystatin C as a GFR marker is that there is no conversion formula transforming cystatin C expressed as mg/L to GFR expressed as mL/min. In this study plasma creatinine and cystatin C were compared with iohexol clearance. A stronger correlation (p < 0.0001) was found between cystatin C and iohexol clearance (r2 = 0.91) than between creatinine and iohexol clearance (r2 = 0.84). From the correlation data a formula was calculated to convert cystatin C expressed as mg/L to GFR (mL/min). The formulas y = 77.24x(-1.2623) (Dade Behring cystatin C calibration) or y = 99.43x(-1.5837) (DakoCytomation cystatin C calibration) are used to calculate GFR expressed in mL/min from the cystatin C value in mg/L and both results are reported to the referral doctor. These formulas can provide the clinicians with reliable and readily available GFR data based on single measurements of cystatin C concentrations.
Innate immunity is important for the integrity of the host against potentially invasive pathogenic microorganisms in the environment. Antibiotic peptides with broad antimicrobial activity are part of the innate immune system. We investigated the presence of the cathelicidin, human cationic antimicrobial protein (hCAP-18), in the male reproductive system. We found strong expression of the hCAP-18 gene by in situ hybridization and hCAP-18 protein, as detected by immunohistochemistry, in the epithelium of the epididymis, but not in the testis. The highest expression in the epididymis was in the caudal part. Western blotting showed a doublet band, the upper part corresponding to the size of hCAP-18 in plasma and neutrophils. Using a specific enzyme-linked immunosorbent assay (ELISA), levels of 86.5 ؎ 37.8 g/ml (mean ؎ standard deviation; range, 41.8 to 142.8 g/ml; n ؍ 10) were detected in seminal plasma from healthy donors, which is 70-fold higher than the level in blood plasma. Flow cytometry and immunocytochemistry revealed the presence of hCAP-18 on spermatozoa. ELISA measurement showed levels of 196 ng/10 6 spermatozoa, corresponding to 6.6 ؋ 10 6 molecules of hCAP-18 per spermatozoon. Our results suggest a key role for hCAP-18 in the antibacterial integrity of the male reproductive system. The attachment of hCAP-18 to spermatozoa may implicate a role for hCAP-18 in conception.The integrity of the human reproductive system against potentially invasive pathogenic microorganisms is crucial. Readily available, preformed antimicrobial proteins of the nonadaptive immune system serve as the body's first line of defense (5), while the adaptive immune system becomes involved if pathogens start to invade. In recent years, several components of the human nonadaptive immune system have been isolated and characterized, among them the only member of the cathelicidins known to exist in humans, the human cationic antimicrobial protein (hCAP-18) (4, 10). This protein is synthesized in neutrophil progenitors in the bone marrow and stored in the specific granules of mature neutrophils (15). It is synthesized as an 18-kDa proprotein from which a 5-kDa C-terminal fragment, LL-37, bearing all of the hitherto known biological activity, is cleaved (8). LL-37 has lipopolysaccharide-binding properties and manifests antibacterial effect against a wide range of bacterial species (11,20). Recently, expression of hCAP-18 has also been demonstrated in the epithelium of several organs, including the vagina, cervix, mouth, and lung (2, 6). The cDNA encoding hCAP-18 has been detected in a library prepared from the human testis, suggesting that the gene is expressed here (1).In the present study, we investigated the presence of hCAP-18 in the male reproductive system. We found expression in the epithelium of the epididymis by in situ hybridization and by immunohistochemistry. High levels of hCAP-18 were found in seminal plasma and in association with spermatozoa, but we were unable to detect expression of the hCAP-18 gene or the presence of hC...
The generation of high quality plasma from whole blood is of major interest for many biomedical analyses and clinical diagnostic methods. However, it has proven to be a major challenge to make use of microfluidic separation devices to process fluids with high cell content, such as whole blood. Here, we report on an acoustophoresis based separation chip that prepares diagnostic plasma from whole blood linked to a clinical application. This acoustic separator has the capacity to sequentially remove enriched blood cells in multiple steps to yield high quality plasma of low cellular content. The generated plasma fulfills the standard requirements (<6.0 x 10(9) erythrocytes/L) recommended by the Council of Europe. Further, we successfully linked the plasmapheresis microchip to our previously developed porous silicon sandwich antibody microarray chip for prostate specific antigen (PSA) detection. PSA was detected by good linearity (R(2) > 0.99) in the generated plasma via fluorescence readout without any signal amplification at clinically relevant levels (0.19-21.8 ng/mL).
BACKGROUND In semen, prostate‐specific antigen (PSA or hK3) digests the gel proteins semenogelin I and II, resulting in liquefaction and the release of motile spermatozoa. We characterized the substrate specificity and zinc‐mediated inhibition of PSA. METHODS The proteolysis of human semenogelin I (SgI) and II (SgII) by PSA was characterized by purification of generated SgI and SgII fragments, N‐terminal sequencing, and mass spectrometry. Zn2+‐inhibition of PSA was studied using a chromogenic substrate. RESULTS Eighteen cleavage sites in SgI and 16 in SgII were identified. Cleavages were identified mainly as the C‐terminal of certain tyrosine and glutamine residues, but also the C‐terminal of histidine, aspartic acid, leucine, serine, and asparagine residues. No cleavages were identified at any arginine, lysine, phenylalanine, tryptophan, or methionine residues, indicating that the substrate specificity of PSA is distinct from that of trypsin, chymotrypsin, tissue kallkrein (hK1), and kallikrein 2 (hK2). Zn2+ ions have a dramatic effect on PSA activity; the data indicate that Zn2+ is a tight‐binding inhibitor of PSA activity. CONCLUSIONS The data will enable the optimized design of PSA activity assays, which may prove instrumental to uncovering the role of PSA in cancer and reproduction. The inhibition data indicate that Zn2+ could regulate PSA activity, which may prove important in the development of efficient inhibitors of PSA activity. Prostate 45:132–139, 2000. © 2000 Wiley‐Liss, Inc.
The plasma concentrations of protein S, protein C and C4b-binding protein (C4BP) were analysed during pregnancy, in the postpartum period and in women using oral contraceptives. Free protein S, measured after precipitation of the C4BP-protein S complexes with 5% PEG 6000, was found to be 8.3 mg/l in the control group, which represents 36.3% of the total plasma protein S content (average 23.5 mg/l). The concentration of protein S was significantly decreased during pregnancy, the lowest levels occurring in the second trimester (14.8 mg/l). The values returned to normal within a few days after delivery. The concentration of free protein S was also decreased, down to an average of 3.7 mg/l at delivery, and did not return to normal within the first week postpartum. The mean concentration of protein S in women using oral contraceptives decreased to 17.7 mg/l and the free fraction went down to 6.6 mg/l. Unlike that of protein S, the plasma concentration of protein C increased during pregnancy, reaching a maximum of 135% in the second trimester. Also, it was significantly higher in the postpartum period and in women using oral contraceptives, than in controls. The level of C4BP was increased throughout pregnancy, with a maximum of 143.4% at delivery. These changes in the plasma levels of proteins C and S during pregnancy indicate that the two proteins differ in the regulation of their synthesis. The major decrease in the level of free protein S may predispose to thrombotic episodes during pregnancy, whereas the increased level of protein C may have the reverse effect. These results indicate the importance of taking into account the normal changes in the plasma levels of protein C and S during pregnancy and the use of oral contraceptives, when evaluating patients with increased risk of thromboembolic disease.
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