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Biomedical subjects

P Stevens

Publications and source records attributed to P Stevens.

At least 109 records · Page 6Linked to original sources
PubMed1980

Detection of candida antigenemia in human invasive candidiasis by a new solid phase radioimmunoassay.

There is a continuing need to develop reliable non-invasive methods to aid in the early diagnosis of disseminatyed candidiasis. We report the development of a solid phase "sandwich" radioimmunoassay (RIA) for the detection of soluble cytoplasmic protein antigens (SPA) of Candida albicans in patients with systemic candidiasis. SPA were prepared by ultrasonic disruption of blastospores of C. albicans. Anti-SPA IgG was covalently linked to a solid phase of polyacrylamide-like microspheres. Standard amounts of patient sample or SPA in pooled normal human serum were reacted with the conjugated microspheres, washed and reacted with anti-SPA IgG-125I. The amount of antigen was directly correlated with the amount of bound 125I-IgG anti-SPA. In a retrospective analysis, circulating SPA in the range of 0.5-1.6 microgram/ml was detected in the serum of 12 of 19 patients (63%) with documented systemic candidiasis. There was no detectable SPA in the serum of 20 hospitalized patients with bacteremia, two patients with aspergillosis, one with cryptococcosis, and three with invasive Candida tropicalis. We observed no crossreaction of the RIA with mannan from C. albicans, coccidioidin or culture filtrate antigen of Aspergillus fumigatus. There was significant inhibition of the RIA by serum samples from two patients with chronic candidiasis suggestive of anti-SPA antibody excess. The solid phase RIA to detect circulating SPA can provide a useful noninvasive method for the diagnosis of systemic C. albicans infection.

Antigens, Fungal↗
PubMed1980

K-1 antigen content and the presence of an additional sialic acid-containing antigen among bacteremic K-1 Escherichia coli: correlation with susceptibility to opsonophagocytosis.

Eighty percent of blood culture isolates of Escherichia coli K-1 are resistant to in vitro opsonophagocytosis by normal human granulocytes and fresh serum. To determine the basis for susceptibility to phagocytosis in 20% of bacteremic K-1 E. coli, we investigated possible quantitative and qualitative immunochemical differences in the K-1 antigen content among resistant and sensitive isolates. We prepared extracts of blood culture K-1 E. coli by sonication and determined the K-1 polysaccharide content per dry weight of bacteria by rocket immunoelectrophoresis using cross-reactive equine anti-group B meningococcal sera. We assessed qualitative differences in the antigen content by crossed immunoelectrophoresis, using an immune globulin fraction and isolated immunoglobulin G (IgG) and IgM from the group B antisera. Three different resistant K-1 isolates contained a mean K-1 content of 48.5 +/- 7.6 mug/mg +/- standard deviation of dry bacteria, and three sensitive isolates contained 23.2 +/- 5.6 mug/mg (P < 0.005). Crossed immunoelectrophoresis of extracts from both sensitive and resistant strains revealed a secondary sialic acid-containing antigen that was electrophoretically different from both the major K-1 antigen and a reference group B meningococcal antigen. This negatively charged secondary antigen was susceptible to Clostridium perfringens neuraminidase degradation and reacted only with IgG whereas the major K-1 antigen reacted only with IgM. This antigen was detected in the extracts of resistant isolates only at 10(10) but not at 10(9) colony-forming units per milliliter. This study demonstrates that (i) the degree of phagocytosis of bacteremic E. coli K-1 isolates is inversely associated with K-1 content, and (ii) more easily phagocytosed (sensitive) K-1 isolates have greater amounts of an additional sialic acid-containing antigen that appears to be unrelated to the previously described O acetyl K-1 antigen.

Antibody Specificity↗
PubMed1979

Relative opsonic and protective activities of antibodies against K1, O and lipid A antigens of Escherichia coli.

The K1 Escherichia coli capsular antigen has been implicated as a virulence factor because of the frequency of isolation of K1 containing strains from certain invasive human infections. In the study of the interaction between K1 strains, normal human polymorphonuclear cells (PMNs) and fresh human serum, we have found varying susceptibility to phagocytosis and killing; thus, the in vitro opsonophagocytic and in vivo protective role of K1, somatic O and core glycolipid antibodies remain unclear. We have therefore examined strains of E. coli with defined susceptibility to phagocytosis by normal PMNs and sera and compared the effect of K1, somatic O and lipid A antibodies in opsonophagocytic tests and mouse protection experiments. K1 E. coli strains demonstrating relative resistance to phagocytosis and killing were effectively opsonized only with specific K1 capsular antisera. Similarly, K1 capsular antisera, but not anti-O or lipid A antisera, also provided protection in mice challenged with a LD100 of K1 E. coli that were "resistant" to phagocytosis. The ability of purified capsular antigens from Neisseria meningitidis group B and K1 "resistant" E. coli to inhibit the phagocytosis of a "sensitive" non-K1 and a K1 E. coli strain of "intermediate" susceptibility to opsonophagocytosis was also investigated. Purified K1 and group B capsular antigens were able to block specific capsular-antibody mediated opsonophagocytosis, yet these capsular antigens failed to inhibit the phagocytosis of non-K1 "sensitive" or K1 "intermediate" E. coli. These studies suggest that K1 antibodies are obligatory for the in vitro and in vivo opsonophagocytosis of "resistant" K1 E. coli and that the K1 antigen must remain in situ on the bacterial surface to exert an anti-phagocytic effect.

Animals↗
PubMed1978

In vitro evaluation of opsonic and cellular granulocyte function by luminol-dependent chemiluminescence: utility in patients with severe neutropenia and cellular deficiency states.

Actively phagocytizing polymorphonuclear leukocytes (PMN) emit light or chemiluminescence (CL) which has been shown to be linked to the oxidative activity of the PMN. The measurement of CL has been demonstrated to be a useful tool for the in vitro assessment of intracellular and opsonophagocytic function of PMN. We have increased the sensitivity of the CL measurement by the addition of luminol to the in vitro reaction of PMN, bacteria, and serum. The presence of luminol, which can be oxidized to emit light, amplifies the detection of CL and PMN cellular activity. This amplification effectively reduces the number of PMN that are necessary for assessment of PMN function from 1 x 10(7) to as low as 2 x 10(4) PMN/assay and permits the evaluation of PMN function in severely neutropenic patients (100 PMN/mm3) in whom cellular PMN function has been heretofore extremely difficult to assess by other methodology. When this luminol-dependent CL method was used, three of eight neutropenic leukemic patients with gram-negative septicemia were found to have deficient opsonic activity and/or increased or depressed cellular oxidative activity. Because the initial slope of CL is dependent on the amount of serum and heat-labile factors, this method can also be used effectively as a simple technique for the analysis of specific rates of opsonophagocytosis of various microorganisms. Additionally, this method can detect the cellular PMN abnormalities of chronic granulomatous disease and myeloperoxidase deficiency. The luminol-dependent CL method is a simple, sensitive, reproducible technique that provides useful information about PMN metabolic activity, particularly in studies in which the number of PMN is limited.

Agranulocytosis↗
PubMed1978

Antibiotic-acrylic bone cement composites. Studies of gentamicin and Palacos.

Palacos bone cement with and without contained gentamicin was tested for antibacterial efficacy using a rabbit model of a knee hemiarthroplasty and a challenge with intra-articular Escherichia coli. The gentamicin-containing cement prevented infection from challenge with Escherichia coli and protected against inadvertent operative infection as well. Antibiotic concentrations in synovial fluid and urine remained in the therapeutic range for three days, whereas levels in serum were always quite low. Eight per cent of the gentamicin had leached out of the bone cement by eight days, most of it during the first day. Thereafter, extremely small amounts of gentamicin left the cement on a daily basis. The amount of antibiotic leaching out in vitro was proportional to the surface area of the cement. More eluted into serum than into saline.

Animals↗
PubMed1977

Cross-protective immunity to Gram-negative bacilli: studies with core glycolipid of Salmonella minnesota and antigens of Streptococcus pneumoniae.

Two immunoprophylactic approaches to the control of infections caused by gramnegative bacilli were evaluated by study of experimental infections in animals. The core glycolipid antigen derived from the Re mutant of Salmonella minnesota R595 is shared by virtually all enteric bacteria, and immunization with this endotoxin protects against the hemodynamic sequelae of bacterial infection and pyrexia without enhancing intravascular clearance of bacteria. The degree of protection afforded by active and passive immunization with core glycolipid was significantly less than that conferred by type-specific immunization. Escherichia coli and Klebsiella pneumoniae share capsular antigens with some strains of Streptococcus pneumoniae; by the mechanism of enhanced opsonization, antibodies to S. pneumoniae may cross-protect against infection with E. coli or K. pneumoniae.

Animals↗
PubMed1977

125I radioimmunoassay of netilmicin.

We report a radioimmunoassay for the new semisynthetic aminoglycoside netilmicin with a sensitivity of 480 pg per tube and a correlation coefficient of 0.94 between this radioimmunoassay and a microbioassay for measurement of netilmicin in serum.

Anti-Bacterial Agents↗
PubMed1977

Quantitative granulocyte chemiluminescence in the rapid detection of impaired opsonization of Escherichia coli.

Previous work has demonstrated that 40% of clinically isolated Escherichia coli are resistant to in vitro killing by normal human polymorphonuclear granulocytes (PMN) due to ineffective opsonophagocytosis. Using these E. coli isolates, we have demonstrated the usefulness of measuring the chemiluminescence (CL) of granulocytes undergoing phagocytosis in detecting this impaired opsonization. CL correlated well with several other methods to assess PMN function including in vitro killing assays, postphagocytic O2 consumption, and morphological phagocytic indexes. Of the available methods to assess granulocyte function, CL is the most rapid and simple and would appear to be a potentially useful screening method to determine possible opsonic deficiencies of human PMNs and serum. Impaired opsonization to these resistant E. coli isolates was demonstrable in several different donors and could be reversed by type-specific rabbit antibody to the particular resistant isolate. Bacterial levels of superoxide dismutase and catalase, enzymes which have been implicated as possible virulence factors in some microorganisms, did not correlate with resistance in these E. coli isolates. However, heat denaturation of these isolates reversed this resistance and would suggest heat-labile antigenic determinants present in E. coli as possible resistance factors.

Blood Bactericidal Activity↗