scholarly journals Development of a Dual Monoclonal Antibody Immunoassay for Total Human Kallikrein 2

2001 ◽  
Vol 47 (7) ◽  
pp. 1218-1224 ◽  
Author(s):  
Judith A Finlay ◽  
John R Day ◽  
Cindy L Evans ◽  
Robert Carlson ◽  
Kristine Kuus-Reichel ◽  
...  
Keyword(s):  
Specific Antigen ◽  
High Specificity ◽  
Cross Reactivity ◽  
Minimum Detectable Concentration ◽  
Serum Samples ◽  
Bodily Fluids ◽  
Before And After ◽  
Detectable Concentration ◽  
Human Kallikrein 2 ◽  
Kallikrein 2

Abstract Background: Human kallikrein 2 (hK2) shares 80% sequence identity with prostate-specific antigen (PSA). Because both hK2 and hK2-α1-antichymotrypsin (hK2-ACT) complexes have been identified in patient sera, we devised an immunoassay for total hK2 [(thK2); hK2 and hK2-ACT] and evaluated it in healthy subjects and patients with prostate disease. Methods: We developed monoclonal antibodies (mAbs) with high specificity for hK2 and hK2-ACT and minimal cross-reactivity to PSA. Using these mAbs, a sandwich assay was developed and its specificity for forms of hK2 was assessed. Serum samples (n = 1035) from healthy volunteers, patients with increased PSA, and men who had undergone radical prostatectomy were assayed for thK2. We also measured thK2 in samples before and after storage under common laboratory conditions. Results: The minimum detectable concentration in the thK2 assay was 0.008 μg/L, and PSA cross-reactivity was <0.001%. The assay detected prohK2 and three different hK2–serum protease complexes. The median serum concentration of thK2 in control samples (0.013 μg/L) was significantly lower than the median in samples from patients with increased PSA concentrations (0.085 μg/L). Immunoreactive hK2 changed little in samples stored for up to 1 month at −70 °C. Conclusions: The thK2 assay recognizes all forms of hK2 that have been found in bodily fluids to date.

scholarly journals Development of Sensitive Immunoassays for Free and Total Human Glandular Kallikrein 2

2004 ◽  
Vol 50 (9) ◽  
pp. 1607-1617 ◽  
Author(s):  
Ville Väisänen ◽  
Susann Eriksson ◽  
Kaisa K Ivaska ◽  
Hans Lilja ◽  
Martti Nurmi ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Detection Limit ◽  
Solid Phase ◽  
Specific Antigen ◽  
Control Group ◽  
Serum Samples ◽  
Human Kallikrein 2 ◽  
Kallikrein 2 ◽  
Capture Antibody ◽  
Total Psa

Abstract Background: Free and total human kallikrein 2 (hK2) might improve the discrimination between prostate cancer and benign prostatic hyperplasia. Concentrations of hK2 are 100-fold lower than concentrations of prostate-specific antigen (PSA); therefore, an hK2 assay must have a low detection limit and good specificity. Methods: PSA- and hK2-specific monoclonal antibodies were used in solid-phase, two-site immunofluorometric assays to detect free and total hK2. The total hK2 assay used PSA-specific antibodies to block nonspecific signal. The capture antibody of the free hK2 assay did not cross-react with PSA. To determine the hK2 concentrations in the male bloodstream, total hK2 was measured in a control group consisting of 426 noncharacterized serum samples. Free and total hK2 were measured in plasma from 103 patients with confirmed prostate cancer. Results: All 426 males in the control group had a total hK2 concentration above the detection limit of 0.0008 μg/L. The median total hK2 concentration was 0.022 μg/L (range, 0.0015–0.37 μg/L). hK2 concentrations were 0.1–58% of total PSA (median, 3.6%). hK2 concentrations were similar in men 41–50 and 51–60 years of age. The ratio of hK2 to PSA steadily decreased from 5–30% at PSA <1 μg/L to 1–2% at higher PSA concentrations. In 103 patients with prostate cancer, the median hK2 concentration in plasma was 0.079 μg/L (range, 0.0015–16.2 μg/L). The median free hK2 concentration was 0.070 (range, 0.005–12.2) μg/L. The proportion of free to total hK2 varied from 17% to 131% (mean, 85%). Conclusions: The wide variation in the free-to-total hK2 ratio suggests that hK2 in blood plasma is not consistently in the free, noncomplexed form in patients with prostate cancer. The new assay is sufficiently sensitive to be used to study the diagnostic accuracies of free and total hK2 for prostate cancer.

scholarly journals Analytical performance of the Tandem®-R free PSA immunoassay measuring free prostate-specific antigen

1997 ◽  
Vol 43 (7) ◽  
pp. 1203-1208 ◽  
Author(s):  
David L Woodrum ◽  
Chester M French ◽  
Timothy M Hill ◽  
Steven J Roman ◽  
Harold L Slatore ◽  
...  
Keyword(s):  
Prostate Specific Antigen ◽  
Accurate Determination ◽  
Specific Antigen ◽  
Cross Reactivity ◽  
Analytical Performance ◽  
Minimum Detectable Concentration ◽  
Free Psa ◽  
Detectable Concentration ◽  
Total Psa

Abstract The analytical performance of the Tandem®-R free PSA assay available from Hybritech Inc. was evaluated. Comparison of recoveries of purified free (unbound) prostate-specific antigen (PSA) diluted in female serum in the Tandem-R free PSA assay and the Tandem-R (total) PSA assay demonstrated a link in calibration between the assays and an accurate determination of percent free PSA. The cross-reactivity of the assay to purified PSA–α1-antichymotrypsin was determined to be <1%. The minimum-detectable concentration was <0.05 μg/L. The within-run and between-day CVs were ≤5% for samples with >0.3 μg/L free PSA. Dilution and recovery showed no significant deviations from linearity across the assay range. The assay was insensitive to interference from blood components. The Tandem-R free PSA kit was shown to be an accurate, precise, and reliable assay for the measurement of free PSA.

Serum Adsorption Study to Validate the Specificity of a Rapid Test to Detect Strongyloides stercoralis Infection

2021 ◽  
Author(s):  
Rahmah Noordin ◽  
Emelia Osman ◽  
Nor Suhada Anuar ◽  
Nor Mustaiqazah Juri ◽  
Anizah Rahumatullah ◽  
...  
Keyword(s):  
Small Sample Size ◽  
Rapid Test ◽  
High Specificity ◽  
Strongyloides Stercoralis ◽  
Cross Reactivity ◽  
Small Sample ◽  
Polystyrene Microspheres ◽  
Diagnostic Specificity ◽  
Serum Samples ◽  
Rapid Tests

A lateral flow rapid test for strongyloidiasis will greatly facilitate the control and elimination of the disease. Previously SsRapid™ prototype rapid test showed high diagnostic specificity to detect Strongyloides infection, determined using non-Strongyloides sera negative by IgG-ELISAs. Since high specificity is crucial before a test is used for public health control activities, further validation of its specificity is needed. Also, it needs to be ascertained whether non-Strongyloides sera positive by IgG-ELISAs and SsRapid are truly positive for Strongyloides or are cases of cross-reactivity. We performed 84 rapid tests (two types of dipsticks and cassettes) using 34 serum samples. They were divided into four groups based on Strongyloides infection and coinfection with other parasites and the availability of recombinant proteins and rapid tests for the latter. Sera was adsorbed using polystyrene microspheres beads separately coated with four recombinant parasite proteins. The small sample size is a limitation of this study; however, the overall results showed that the sera adsorption procedure was successful, and the SsRapid test is specific.

scholarly journals Immunofluorometric Assay of Human Kallikrein 10 and Its Identification in Biological Fluids and Tissues

2001 ◽  
Vol 47 (2) ◽  
pp. 237-246 ◽  
Author(s):  
Liu-Ying Luo ◽  
Linda Grass ◽  
David J C Howarth ◽  
Pierre Thibault ◽  
Huy Ong ◽  
...  
Keyword(s):  
Biological Fluids ◽  
Expression System ◽  
Specific Antigen ◽  
Cross Reactivity ◽  
Yeast Expression System ◽  
Disease States ◽  
Pichia Pastoris Yeast ◽  
Sandwich Type ◽  
Kallikrein 2 ◽  
Concentration Changes

Abstract Background: The human kallikrein 10 gene [KLK10, also known as normal epithelial cell-specific 1 gene (NES1)] is a member of the human kallikrein gene family. The KLK10 gene encodes for a secreted serine protease (hK10). We hypothesize that hK10 is secreted into various biological fluids and that its concentration changes in some disease states. The aim of this study was to develop a sensitive and specific immunoassay for hK10. Methods: Recombinant hK10 protein was produced and purified using a Pichia pastoris yeast expression system. The protein was used as an immunogen to generate mouse and rabbit polyclonal anti-hK10 antisera. A sandwich-type immunofluorometric assay was then developed using these antibodies. Results: The hK10 immunoassay has a detection limit of 0.05 μg/L. The assay is specific for hK10 and has no detectable cross-reactivity with other homologous kallikrein proteins, such as prostate-specific antigen (hK3), human glandular kallikrein 2 (hK2), and human kallikrein 6 (hK6). The assay was linear from 0 to 20 μg/L with within- and between-run CVs <10%. hK10 is expressed in many tissues, including the salivary glands, skin, and colon and is also detectable in biological fluids, including breast milk, seminal plasma, cerebrospinal fluid, amniotic fluid, and serum. Conclusions: We report development of the first immunofluorometric assay for hK10 and describe the distribution of hK10 in biological fluids and tissue extracts. This assay can be used to examine the value of hK10 as a disease biomarker.

The value of (−7, −5)pro-prostate-specific antigen and human kallikrein-2 as serum markers for grading prostate cancer

2004 ◽  
Vol 93 (6) ◽  
pp. 720-724 ◽  
Author(s):  
C.H. Bangma ◽  
M.F. Wildhagen ◽  
G. Yurdakul ◽  
F.H. Schröder ◽  
B.G. Blijenberg
Keyword(s):  
Prostate Cancer ◽  
Prostate Specific Antigen ◽  
Specific Antigen ◽  
Serum Markers ◽  
Human Kallikrein 2 ◽  
Kallikrein 2

scholarly journals Benign Prostate-specific Antigen (BPSA) in Serum Is Increased in Benign Prostate Disease

2003 ◽  
Vol 49 (2) ◽  
pp. 253-259 ◽  
Author(s):  
Harry J Linton ◽  
Leonard S Marks ◽  
Lisa S Millar ◽  
Christine L Knott ◽  
Harry G Rittenhouse ◽  
...  
Keyword(s):  
Prostate Specific Antigen ◽  
Limit Of Detection ◽  
Specific Antigen ◽  
Clinical Information ◽  
High Specificity ◽  
Wide Distribution ◽  
Cross Reactivity ◽  
Free Psa ◽  
Control Serum ◽  
Benign Prostate

Abstract Background: BPSA is a “benign” form of free prostate-specific antigen (PSA) that is increased in prostate transition zone tissues of men with pathologic benign prostatic hyperplasia (BPH). We developed an immunoassay to determine the concentration of BPSA in the serum of men with BPH. Methods: The BPSA antigen was purified by HPLC, and murine monoclonal antibodies were prepared by standard methods. A fluorogenic ELISA was developed with high specificity for BPSA and no cross-reactivity with other forms of PSA. Results: The BPSA immunoassay had a lower limit of detection of 6 ng/L and a cross-reactivity of <1% with all other clipped and nonclipped forms of PSA. The BPSA antibody was specific for the internal Lys182 cleavage site that characterizes BPSA. Biopsy-negative men with a median total PSA of 4.8 μg/L had a median of 0.22 μg/L BPSA, representing 25% of the free PSA in serum. BPSA ranged from 0% to 60% of the free PSA in serum. BPSA in a cohort of cancer serum also comprised 25% of the free PSA. Control serum from women or men without increased PSA had nondetectable BPSA. Conclusions: BPSA is a significant percentage of the free PSA in BPH serum but not in control serum. The presence of prostate cancer does not alter the relative proportions of BPSA in sera with <10 μg/L PSA. BPSA has a wide distribution of concentrations in the serum and may provide clinical information for the study of men with BPH.

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