SEARCH · PubMed Health

Results for “ELISA assay variability”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 145 records · Page 8Linked to original sources

PubMed2001

Correlation between serum hyaluronan and disease activity and severity in Thai patients with rheumatoid arthritis.

Serum hyaluronan (HA) concentration was quantified using an ELISA-based assay with biotinylated hyaluronan binding proteins, and correlated with the clinical and laboratory variables in 100 consecutive rheumatoid arthritis (RA) patients (mean +/- SD age and duration of disease of 50.1 +/- 12.5 and 7.9 +/- 6.6 years respectively). Thirty-four patients received prednisonole at an average dose of 5.0 mg/day. The correlations were good between the serum HA level and the joint swollen scores (r = 0.26, p = 0.04), joint space narrowing scores (r = 0.25, p = 0.03), joint erosion scores (r = 0.26, p = 0.03), and erythrocyte sedimentation rate (r = 0.31, p < 0.01) in RA patients who did not take prednisolone. These correlations were diminished in those who received prednisolone, although their disease was more severe. It might be possible that corticosteroids could decrease inflammation of the joint, thus interfering with the correlations. It was concluded that the serum HA level is a useful marker for the activity and severity of disease in patients with RA.

Adult↗

PubMed1995

[Determination of phenobarbital by solid-phase immunoenzyme analysis. Choice of optimal strategy].

Two variants (direct and indirect) of enzyme linked immunosorbent assay (ELISA) of phenobarbital are compared. Both techniques were developed on the basis of the same monoclonal antibodies, and horse radish peroxidase was used as the label in both cases. When microtitration plates are used as the solid phase, indirect ELISA, in which phenobarbital of the sample competes with phenobarbital sorbed on plates in the form of a conjugate with protein for the binding with peroxidase-labeled antiphenobarbital antibodies, is preferable. In indirect ELISA, the sample volume was 5 microliters, the time of assay was 40 min, the variability coefficient was < 8%.

Animals↗

PubMed2001

Increased level of YKL-40 in sera from patients with early rheumatoid arthritis: a new marker for disease activity.

YKL-40 is a newly discovered major secretory protein of human chondrocytes and synoviocytes. We measured serum levels of YKL-40 in 52 patients with early onset rheumatoid arthritis (RA) by enzyme-linked immunosorbent assay (ELISA) during a 2-year prospective follow-up, correlating values with laboratory and clinical variables and radiographic progression. Levels at baseline before antirheumatic therapy were significantly higher in patients than in healthy controls. The levels of YKL-40 correlated with laboratory and clinical markers of disease activity both at baseline and during follow-up. Baseline YKL-40 values correlated with baseline Larsen scores but did not predict radiographic progression. Baseline and mean YKL-40 values did not differ between fast and slow radiological progressions. Mean YKL-40 levels correlated with the number of swollen joints but were not predictors of radiographic progression. These results suggest that in early RA, serum YKL-40 is an inflammatory marker correlating with disease activity. However, its levels do not predict clinical course or radiographic progression.

Adipokines↗

PubMed2005

Quantification of thrombin activatable fibrinolysis inhibitor (TAFI) gene polymorphism effects on plasma levels of TAFI measured with assays insensitive to isoform-dependent artefact.

Reports have reappraised the genotype-dependent variation of Thrombin Activatable Fibrinolysis Inhibitor (TAFI), demonstrating that, in some enzyme-linked immunosorbent assays (ELISA), decreased antibody reactivity towards the TAFI 325 Ile isoform led to erroneous TAFI levels. Assays free of this artefact are required to evaluate the contribution of the TAFI gene polymorphisms to TAFI level variability. TAFI levels were measured in 209 individuals with both immunological and functional assays. Each assay was characterized, in particular for its reactivity towards the 325 Thr and Ile isoforms. Two ELISAs were found to have a lower reactivity towards the Ile isoform, leading to an overestimate of the magnitude of variation between two different genotypes. In contrast, one ELISA, as well as functional assays, was found to be free of genotype-dependent artefact constituting a reliable method to detect true quantitative variations of TAFI levels. Using these reliable methods, univariate and haplotype analyses revealed that TAFI gene polymorphisms explain 25% of TAFI level variability. This effect seems to be the consequence of the joint independent action of two polymorphisms, one in the 5' (G-1102T) and the other in the 3' region (T+1583A) of the TAFI gene.

Alleles↗

PubMed1994

Analysis of antibody specificity against the third variable region of the envelope glycoprotein gp120 of HIV-1 in plasma from HIV-1-positive individuals residing in Brazil.

1. Antibody specificity for the principal neutralization domain (PND) of the human immunodeficiency virus type 1 (HIV-1) was studied in plasma from 122 HIV-1-infected individuals residing in Brazil. 2. Using 8 overlapping sequential pentadecapeptides corresponding to the third variable region (V3) of 5 different HIV-1 isolates in an enzyme-linked immunosorbent assay (ELISA), a preferential recognition of the peptides with amino acid sequences corresponding to the HIV-1 isolates IIIB and MN (maximal reactivities of 60-70%) compared to the isolates SC, WMJ-2 or RF (maximal reactivities below 60%) was observed. 3. A difference was observed in the overall reactivity pattern to HIV-1 SC peptides of plasma collected from individuals residing in the Brazilian states of Rio de Janeiro and Bahia. However, a statistically significant increased recognition by Bahian plasma was only observed for the HIV-1 SC C55 peptide. 4. The mean CD4/CD8 ratio of the group of plasma with an isolate-restricted recognition of peptides (0.522 +/- 0.074) was significantly lower than that of the total group of plasma (1.00 +/- 0.18).

Amino Acid Sequence↗

PubMed1997

Interlaboratory agreement among results of human papillomavirus type 16 enzyme-linked immunosorbent assays.

Serological assays for measuring antibodies to human papillomavirus type 16 (HPV-16) virus-like particles (VLPs) have become important epidemiologic tools in recent years. However, the interlaboratory replicability of these assays has not been assessed. In this investigation, three laboratories tested a panel of specimens obtained from two different groups: 265 subjects in a vulvar cancer case-control study and 107 healthy volunteer blood donors. Each laboratory used an enzyme-linked immunosorbent assay (ELISA), but no attempt was made to standardize assay procedures among the three laboratories. The data showed good day-to-day intralaboratory replicability in laboratory 1 (correlation coefficient, > or = 0.88) and good intra-assay variability in laboratory 3 (correlation coefficient, > or = 0.93). Interlaboratory correlations, likewise, ranged between 0.61 and 0.80 in both case-control study subjects and healthy blood donors, indicating that ELISA optical density (OD) values between laboratories were linearly related regardless of the population. Kappa coefficients (kappa), based on each laboratory's categorical interpretation of its results (as positive or negative), showed good agreement (kappa, > 0.6) in case-control study subjects and moderate agreement (kappa, > or = 0.4) in blood donors, a population that had few strongly positive sera. When OD values near seropositive cutoffs were treated as indeterminates, there was little discordance between laboratories in either population. The data suggest that each laboratory measured the same humoral immune response and that their HPV-16 VLP ELISAs performed similarly (Pearson correlations). Interlaboratory differences, however, probably due to reagents and procedures, were considerably greater than intralaboratory day-to-day variability. Interlaboratory agreement in determining seropositivity (kappa) could be improved by sharing positive and negative serum controls and by treating marginal results as indeterminate. As part of continuing cooperation to improve interlaboratory agreement, we are preparing bulk serum control specimens to be shared and made available to interested researchers.

Enzyme-Linked Immunosorbent Assay↗

PubMed1994

Antihistone antibodies in systemic lupus erythematosus: assay dependency and effects of ubiquitination and serum DNA.

OBJECTIVE: Antihistone antibodies occur in systemic lupus erythematosus (SLE) but there are many discrepancies in their reported prevalence, isotype, specificity and correlation with disease symptoms. We examined the role of the assay and the influence of serum DNA as possible causes of these discrepancies. In addition, we sought to confirm the presence of antibodies to ubiquitin and ubiquitinated H2A (uH2A). METHODS: Western blot and enzyme linked immunosorbent assay (ELISA). RESULTS: Sera displayed substantial differences between ELISA and Western blot in reactivity to individual histones when all reagents were nearly identical, indicating that subtle differences in the solid phase adsorbents have pronounced effect on histone antigenicity. No uniform pattern of antibody reactivity with the 5 histones was apparent with either assay. For most sera, digestion with DNase caused only minor decrease in binding to histones and no histone class showed particular sensitivity to this treatment. In agreement with most other studies, no significant correlation between histone binding and symptoms was found. Just 2 of 40 sera showed detectable binding to ubiquitin or uH2A. CONCLUSION: Although IgG antihistone antibodies were detected in 53-55% of patients with SLE with active disease, the sensitivity of antibody activity to assay conditions, patient variability, and lack of correlation with symptoms compromise the clinical utility of measuring antihistone antibodies by Western blot or ELISA: We were also unable to confirm that ubiquitin and uH2A are major antigens recognized by antibodies in SLE.

Adult↗

PubMed2000

Open sandwich enzyme-linked immunosorbent assay for the quantitation of small haptens.

The quantitation of low-molecular-weight haptens has been difficult with conventional sandwich immunoassays due to their small size. Many researchers have attempted to develop sandwich assays for haptens due to the significant advantages of the sandwich format over competitive assays including greater dynamic range, ease of automation, and sensitivity. Here we apply the open-sandwich ELISA (OS-ELISA), an immunoassay based on antigen-dependent stabilization of antibody variable regions (V(H) and V(L) domains), to hapten quantitation. Two fusion proteins, the high-affinity mutant V(H) domain from anti-4-hydroxy-3-nitrophenacetyl (NP) antibody B1-8 tethered with Escherichia coli alkaline phosphatase (V(H)(W33L)-PhoA) and the V(L) domain from the same antibody tethered with Streptococcus sp. protein G, were made. These fusion proteins when added together achieved Fv reassociation consequent to the addition of NP. Signal was generated in a direct relationship to the NP concentration with better sensitivity compared with competitive immunoassay, demonstrating this assay to be a quick noncompetitive alternative to the conventional assays for small compounds, such as environmental pollutants, drugs of abuse, and therapeutic drugs. With our previous demonstration that the OS-ELISA works well with large proteins, the OS-ELISA becomes the first practical immunoassay approach capable of quantifying any molecule regardless of their size.

Animals↗

PubMed1991

Comparative evaluation of 36 commercial assays for detecting antibodies to HIV.

Summarized are the results of an assessment of the major operational characteristics of 36 commercially available assays for detection of antibodies to human immunodeficiency virus type 1 (HIV-1) and/or type 2 (HIV-2). For this purpose, 20 enzyme-linked immunosorbent assays (ELISAs), 11 simple immunoassays with visual reading, four supplemental assays, and one discriminatory assay were assessed using a panel of 537 sera (65% of which were of African, 26% of European, and 9% of South American origin); the prevalence of HIV-1 was 39.1% and of HIV-2, 15.7%. The following operational parameters of the assays were investigated: ease of performance; suitability for use in small blood collection centres; sensitivity and specificity; positive predictive values at different prevalences; inter-reader variability for simple assays whose results were read visually; the proportion of indeterminate results; and, for some of the ELISA assays, delta-values, as quantitative measures of sensitivity and specificity. The results will be of use to health policy decision-makers, managers of national AIDS prevention and control programmes, directors of blood banks, and laboratory specialists in the selection of appropriate HIV antibody assays.

AIDS Serodiagnosis↗

PubMed1991

Immunochemical characterization of variable epitopes of outer membrane protein P2 of nontypeable Haemophilus influenzae.

Monoclonal antibodies (MAbs) were elicited to the nontypeable Haemophilus influenzae variants d1 to d4, which differ in the outer membrane protein P2 to analyze the immunological properties of the variable parts of this protein. Five MAbs reacted in a whole-cell enzyme-linked immunosorbent assay (ELISA) only with the homologous strain and in some cases with its variants, but not with 69 unrelated nonencapsulated H. influenzae isolates; nine MAbs also reacted with some other H. influenzae isolates, and four MAbs showed broad cross-reactivity. All of the MAbs reacted with purified protein P2 in ELISAs and immunoblotting. The five MAbs which reacted with the homologous strain d3 and not with the variants d1, d2, and d4 promoted complement-dependent bactericidal activity against strain d3. These and four other MAbs reacted with the intact bacteria of strain d3 in immunogold electron microscopy, indicating that they were directed against surface-exposed epitopes of outer membrane protein P2. A mutant of strain d3 was isolated as a survivor from bacterial killing by complement and MAb 30DA5. This mutant had an altered P2 protein on sodium dodecyl sulfate-polyacrylamide gels and had lost its reactivity with all of the five H. influenzae d3-specific MAbs but not with the other MAbs. From these results, we conclude that the variable parts of outer membrane protein P2 of nonencapsulated H. influenzae from the sputum of patients with chronic obstructive pulmonary disease are immunogenic and mostly surface exposed. Only strain-specific MAbs promoted complement-dependent killing of the bacteria, which was abolished in a spontaneous mutant with an altered P2 protein.

Animals↗

PubMed1994

Adjuvant effect of pertussis toxin on the production of anti-ovalbumin IgE in mice and lack of direct correlation between PCA and ELISA.

In studies designed to optimize the production and detection of anti-ovalbumin (anti-Oa) IgE in Ham/ICR mice, a range of doses of both Oa and pertussis toxin as the IgE adjuvant was explored. As determined by 48-hour passive cutaneous anaphylaxis (PCA) tests, the highest titre of anti-Oa IgE was obtained in a three-injection protocol with 0.1 micrograms Oa and 1 microgram pertussis toxin for the priming dose, followed by two further doses of 0.1 microgram Oa alone. In single-dose immunizations, the highest PCA responses were obtained in sera from mice given 20 micrograms Oa and 1 microgram pertussis toxin. These data confirm that murine IgE production to Oa depends on particular combinations of immunization variables. There was no direct correlation between the PCA and anti-IgE enzyme-linked immunosorbent assay (ELISA) titres of the PCA-positive sera; indeed there was a significant negative correlation. This relationship was not due to interference by IgG1, as there was no correlation between the anti-Oa IgG1 and anti-Oa IgE ELISA titres of the sera. These results highlight the need for caution in assuming that serum IgE levels as measured by ELISA will necessarily correlate positively with IgE biological activity as measured by allergen challenge in vivo.

Adjuvants, Immunologic↗

PubMed2005

Two immunochemical assays to measure advanced glycation end-products in serum from dialysis patients.

Advanced glycation end-products are uremic toxins that accumulate in the serum and tissues of patients with chronic renal failure. Here, we established two enzyme-linked immunosorbent assays (ELISAs) for N(epsilon)-carboxymethyllysine and imidazolone to analyze advanced glycation end-products in human serum. Both ELISAs detected advanced glycation end-products bound to human serum albumin in a dose-dependent way. Whereas the formation of imida-zolone was independent of the presence of oxygen, concentrations of N(epsilon)-carboxymethyllysine epitopes increased 20-fold under oxidative conditions. The N(epsilon)-carboxymethyllysine ELISA showed a similar response to free, peptide-bound and protein-bound N(epsilon)-carboxymethyllysine, whereas the imidazolone antibody showed slightly higher affinity toward peptide-bound compared to protein-bound imidazolone. In human serum, linear dilution ranges from 1:10 to 1:40 (N(epsilon)-carboxymethyllysine ELISA) and from 1:2 to 1:8 (imidazolone ELISA) were found. The recovery of N(epsilon)-carboxymethyllysine from serum was 101 +/- 10% and 94 +/- 12%, respectively, and 93 +/- 15% and 97 +/- 12% for imidazolone. The coefficients of variation for intra-assay variability were 0.26-2.7% (N(epsilon)-carboxymethyllysine) and 0.1-2.4% (imidazolone), and 8.3-13.4% (N(epsilon)-carboxymethyllysine) and 7.8-12.5% (imidazolone) for inter-assay variability. In serum samples from hemodialysis patients (n = 20) and controls (n =20), an approximately two-fold increase was detected in the patient group (p < 0.001). The combination of the N(epsilon)-carboxymethyllysine and imidazolone ELISAs is a valuable tool to measure serum concentrations of advanced glycation end-products for clinical studies.

Adult↗

PubMed2005

The Study of the Effects of Diet on Metabolism and Nutrition (STEDMAN) weight loss project: Rationale and design.

This paper outlines the rationale and design of the Study of the Effects of Diet on Metabolism and Nutrition (STEDMAN) weight loss project, in which detailed biologic profiling of three hundred and fifty obese individuals (body mass index (BMI): 30-50 kg/m(2)) will be conducted as they lose weight via seven distinct interventions. These profiles will be compared to those of fifty normal, healthy, control participants (BMI: 18.5-24.9 kg/m(2)). The interventions include the following: Roux-en-Y gastric bypass surgery, dietary interventions of differing macronutrient composition and diverse pharmacologic interventions. Outcome variables include eight conventional metabolites and CRP measured by standard clinical chemistry techniques, twenty hormones of energy balance and fuel homeostasis measured by radioimmunoassay (RIA) or by enzyme-linked Immunosorbent assay (ELISA), ten pro- and anti-inflammatory cytokines measured using Luminex xMAP technology, one hundred and one intermediary metabolites measured by targeted mass-spectrometry-based methods, and physiologic variables such as body composition measured by dual energy X-ray absorptiometry (DEXA), air displacement plethysmography, and abdominal computerized tomography (CT), insulin sensitivity measured by intravenous glucose tolerance test (IV-GTT) and metabolic rate measured by indirect calorimetry. Results from this study will expand our knowledge of the biology of obesity and weight regulation and may lead to targeted strategies for its treatment and control.

Adolescent↗

PubMed2006

ELISA methods detect HLA antibodies with variable sensitivity.

The accuracy of human leukocyte antigen (HLA) antibody screening results is dependent on the technique employed. In this study, a total of 612 serum samples from patients awaiting kidney transplantation were tested by two different enzyme-linked immunosorbent assay (ELISA) methods: LAT-M(R) (One Lambda/BMT) and AbScreen(R) (Biotest). The results of these assays were identical for HLA class I and II antibodies in 524 cases (85.6%), and discrepant in the remaining 88 cases (42 class I, 24 class II, 22 class I and II). More specifically, class I results were in agreement in 26 positive and 522 negative cases and in disagreement in 64 cases. Class II results were the same in 50 positive and 516 negative cases and different in 46 cases. Retesting the samples using LAT 1288 (One Lambda/BMT) and considering previous HLA antibody test results and the history of immunising events, the sensitivity and specificity, respectively, of the two methods were determined as follows: class I: 89.6% and 97.5% (LAT-M) vs. 70.6% and 96.8% (AbScreen), class II: 96.5% and 98.4% (LAT-M) vs. 88.3% and 95.5% (AbScreen). These results indicate that a single ELISA does not invariably prove or exclude the presence of HLA antibodies, and that additional testing is required in some cases.

Antibody Specificity↗

PubMed1996

Enzyme-linked immunosorbent assay detects a potential soluble form of the erythropoietin receptor in human plasma.

The erythropoietin receptor (EpoR) is a type I transmembrane protein that is a member of the family of hemopoietin receptors. Several members of this family have soluble receptor forms that are secreted by the cells rather than expressed on the cell surface. An alternatively spliced EpoR transcript has been described in human erythroid precursors that, if translated, would produce a truncated, soluble EpoR lacking the transmembrane domain. To determine if the human EpoR is expressed in a soluble form, we developed a sensitive enzyme-linked immunosorbent assay (ELISA) for the EpoR, and we analyzed human serum and plasma. Sheep were immunized with a fusion protein (EREx) consisting of glutathione-S-transferase (GST) and the human EpoR extracellular domain. The sheep antiserum was affinity-purified on immobilized EREx, and then used in a two-stage antigen capture ELISA. The plasma from 20 normal subjects was studied with this assay. There was a wide variability in the levels of soluble EpoR in these subjects (range, <10-2,200 ng/ml). An average value of 550 +/- 735 ng/ml for soluble EpoR was obtained in these normals. Protein A adsorption of the test plasma prior to the assay had no effect on the values obtained. Assay of serum from the same normal subjects showed an average decrease of 88% in soluble EpoR levels compared to plasma. There was no correlation between hematocrit and soluble EpoR levels compared to plasma. There was no correlation between hematocrit and soluble EpoR level. This assay may have utility in the further elucidation of erythropoietin physiology.

Adult↗

PubMed2003

The relationship of FcgammaRIIIa genotype to degree of B cell depletion by rituximab in the treatment of systemic lupus erythematosus.

OBJECTIVE: Despite wide use of the anti-CD20 monoclonal antibody rituximab in the treatment of B cell lymphomas, the mechanism by which it causes B cell depletion remains a subject of controversy. As part of an ongoing phase I/II trial of rituximab in the treatment of systemic lupus erythematosus (SLE), we sought to determine whether the effectiveness of B cell depletion was influenced by polymorphisms of Fc receptors (FcR) on effector cells. METHODS: During rituximab treatment of 12 SLE patients, B cell depletion was monitored as a function of the serum rituximab level and FcgammaRIIa and FcgammaRIIIa genotypes at baseline and at 1 month and 2 months after treatment. FcR genotypes were determined by polymerase chain reaction. Serum levels of rituximab were measured by enzyme-linked immunosorbent assay (ELISA). B lymphocyte percentages were assessed by flow cytometry. RESULTS: B cell depletion was highly variable in this patient cohort, with B cell percentages at the 1-2-month posttreatment nadir ranging from undetectable (<0.1 cell/microl) to 16% ( approximately 30 cells/microl) of the total peripheral blood lymphocytes. At 2 months posttreatment, B cell percentages were highly correlated with both the serum rituximab level and the FcgammaRIIIa genotype (R(2) = 0.75, P = 0.002). The FcgammaRIIIa genotype was a significant independent predictor of the efficacy of B cell depletion (P = 0.019). CONCLUSION: These results highlight the potential variability of B cell depletion by rituximab in the treatment of autoimmune disease and indicate that Fc receptors are an important determinant of that variability. The findings further suggest the importance of antibody-dependent cell-mediated cytotoxicity and/or apoptosis induction via FcgammaRIIIa-expressing effector cells in the mechanism of B cell depletion by this widely used monoclonal antibody.

Antibodies, Monoclonal↗

PubMed1995

Multiple molecular forms of pyridinolines cross-links excreted in human urine evaluated by chromatographic and immunoassay methods.

The measurement of the collagen cross-links, hydroxylysylpyridinoline (HP) and lysylpyridinoline (LP), excreted in urine either in free or peptide-bound forms represents the most extensively investigated biochemical marker of bone collagen degradation. We studied the urinary molecular forms of pyridinolines after separation in free and peptide-linked fractions by chromatography and serial dialysis. The pyridinoline amounts of molecular species (free, < 1000 D, 1000-3500 D, 3500-10,000 D, and > 10,000 D) were evaluated by high performance liquid chromatography (HPLC) as well as with the two newly introduced enzyme-linked immunosorbent assay (ELISA) methods for determination of free pyridinolines (collagen Pyrilinks and collagen Pyrilinks-D). The variability of urinary pyridinoline forms were studied in healthy adult control subjects (n = 10, 38.4 +/- 7.5) years), in adolescents (n = 10, 16 +/- 3.3 years), and in elderly subjects with vitamin D insufficiency (n = 10, 87.3 +/- 4.3 years). Free and peptide-conjugated pyridinolines with MW < 1000 D constitute the major part of urinary cross-links in all groups, with a significantly lesser excretion in elderly patients than in adolescent groups. Expressed as a percent of total cross-links, urinary free pyridinolines assessed by direct HPLC are less in elderly subjects (HP = 34.2 +/- 6.2%, LP = 32.7 +/- 7.6%) than in adolescents (HP = 45.8 +/- 10.8%, p = 0.0065 and LP = 47.8 +/- 12.1%, p = 0.012) and in healthy adults (HP = 39.3 +/- 11.5%, NS and LP = 38.1 +/- 9.3%, NS).(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗

PubMed2004

The interaction of streptococcal inhibitor of complement (SIC) and its proteolytic fragments with the human beta defensins.

Streptococcal inhibitor of complement (SIC) is a 31 kDa extracellular protein produced by a few highly virulent strains of Streptococcus pyogenes (in particular the M1 strain). It has been shown additionally to inhibit four further components of the mucosal innate response-lysozyme, secretory leucocyte proteinase inhibitor, human alpha-defensin 1 and the cathelicidin LL-37 which are all bactericidal against Group A Streptococci (GAS). We now show that SIC also inhibits variably the antibacterial action of hBD-1, -2 and -3. By enzyme-linked immunosorbent assay (ELISA), SIC binds strongly to hBD-2 and hBD-3, but not at all to hBD-1. Investigation of the antimicrobial action of beta-defensins hBD-1, -2 and -3 against GAS in two different buffer systems shows that both the killing efficiencies of all three defensins, and the binding of SIC to them, occurs more efficiently in 10 mm Tris buffer than in 10 mm phosphate. The lower ionic strength of the Tris buffer may underlie this effect. hBD-1 kills the M1 strain of GAS only in 10 mm Tris, but is able to kill an M6 (SIC negative) strain in 10 mm phosphate. The inhibition of hBD-3 by SIC is clearly of physiological relevance, that of hBD-2 is likely to be so, but the inhibition of hBD-1 occurs only at lower ionic strength than is likely to be encountered in vivo. Elastase digestion of SIC yields three major fragments of MW 3.843 kDa comprising residues 1-33 (fragment A); 10.369 kDa comprising residues 34-126 (fragment B); and MW 16.487 kDa, comprising residues 127-273 (fragment C). By ELISA, only fragment B binds to hBD-2 and hBD-3 and this may indicate the inhibitory portion of the SIC molecule.

Antimicrobial Cationic Peptides↗