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Biomedical subjects

K O Smith

Publications and source records attributed to K O Smith.

At least 19 recordsLinked to original sources
PubMed1991

Enzyme-linked immunosorbent assay (ELISA) for HIV antibody by a glass slide technique.

An enzyme-linked immunosorbent assay (ELISA) technique is described which utilizes a commercially available glass microscope slide coated with hydrophobic teflon in such a pattern as to give 30 small circular wells, each of which has a glass bottom. Each well serves as a solid phase, analogous to a microtiter well for adsorption of purified human immunodeficiency virus (HIV) antigens. Since only 5-10 microliter volumes of reagents are used and rinsing processing is simple, the cost per test is much less than most other ELISA technologies. HIV antigen is stable for over 1 year at 37 degrees C when dried on the glass slides. The sensitivity and specificity of the micro slide immunoenzymatic assay (Micro-SIA) was studied by testing randomly selected, known HIV-seropositive and seronegative plasma. Results compare well with microtiter and Western blot assays. A simple vertical-beam colorimeter is described (useful in the Micro-SIA) which can be easily assembled by the user from commonly available components.

Enzyme-Linked Immunosorbent Assay↗
PubMed1991

Enhanced replication of herpes simplex virus type 1 in human cells.

The effects of DNA-damaging agents on the replication of herpes simplex virus type 1 (HSV-1) were assessed in vitro. Monolayers of human lung fibroblast cell lines were exposed to DNA-damaging agents (methyl methanesulfonate [MMS], methyl methanethiosulfonate [MMTS], ultraviolet light [UV], or gamma radiation [GR]) at specific intervals, before or after inoculation with low levels of HSV-1. The ability of cell monolayers to support HSV-1 replication was measured by direct plaque assay and was compared with that of untreated control samples. In this system, monolayers of different cell lines infected with identical HSV-1 strains demonstrated dissimilar levels of recovery of the infectious virus. Exposure of DNA-repair-competent cell cultures to DNA-damaging agents produced time-dependent enhanced virus replication. Treatment with agent before virus inoculation significantly (p less than 0.025) increased the number of plaques by 10 to 68%, compared with untreated control cultures, while treatment with agent after virus adsorption significantly increased (p less than 0.025) the number of plaques by 7 to 15%. In a parallel series of experiments, cells deficient in DNA repair (xeroderma pigmentosum) failed to support enhanced virus replication. These results suggest that after exposure to DNA-damaging agents, fibroblasts competent in DNA repair amplify the replication of HSV-1, and that DNA-repair mechanisms that act on a variety of chromosomal lesions may be involved in the repair and biological activation of HSV-1 genomes.

Cell Line↗
PubMed1984

Experimental ocular herpetic infections in rabbits. Treatment with 9-([2-hydroxy-1-(hydroxymethyl)ethoxy]methyl)guanine.

A new acyclic nucleoside, 9-([2-hydroxy-1-(hydroxymethyl)ethoxy]methyl)guanine (BIOLF-62), was found to be efficacious in the treatment of experimental ocular herpes simplex virus infections in rabbits. Complete healing of herpetic lesions occurred in a majority of animals after six days of topical treatment, three times per day. No toxic effects were observed in uninfected, drug-treated eyes. The BIOLF-62 treatment blocked viral replication in infected eyes sufficiently to make recovery of virus from any of the drug-treated eyes impossible, whereas virus was recovered from all placebo-treated eyes.

Acyclovir↗
PubMed1984

A diagnostic-prognostic test for bladder cancer using a monoclonal antibody-based enzyme-linked immunoassay for detection of urinary fibrin(ogen) degradation products.

An enzyme-linked immunosorbent assay (ELISA) using a monoclonal antibody was developed to determine the clinical value of urinary fibrinogen/fibrin degradation product levels for the identification and management of patients with bladder cancer. Assays were performed on 286 serial urine specimens from 56 bladder carcinoma patients. Specimens were grouped according to whether the patient had an evident tumor at the time of specimen collection (134 specimens, 41 patients) or was clinically disease-free following treatment (152 specimens, 38 patients). Many patients contributed specimens to both groups as determined by their clinical status at the time of collection. In addition, 45 specimens from 33 patients with inflammation of the urogenital tract and 81 specimens from 19 patients with renal or prostatic cancer were assayed for urinary fibrin degradation products. The ELISA, using a high-sensitivity procedure, identified 83% of the specimens from bladder cancer-positive patients with an overall accuracy with all specimens of 78% and a false-negative rate of 5% for all specimens tested. The high-sensitivity ELISA appeared most appropriate for monitoring bladder cancer patients for recurrence of tumor after surgery. The ELISA using a high-specificity procedure appeared most appropriate for screening. The high-specificity ELISA accurately identified 96% of urine specimens from non-bladder cancer patients with a false-positive rate of only 5%. These results demonstrate that the ELISA is an efficient, reliable, quantitative, and noninvasive immunoassay that can be useful both for the identification of bladder cancer patients and for monitoring the course of the disease.

Aged↗
PubMed1983

Sensitivity of equine herpesviruses 1 and 3 in vitro to a new nucleoside analogue, 9-[[2-hydroxy-1-(hydroxymethyl) ethoxy] methyl] guanine.

The following members of the Herpetoviridae family were tested to determine their sensitivities to the new antiviral drug, BIOLF-62: equine herpesvirus types 1 and 3, human herpesvirus types 1 and 2, swine herpesvirus, bovine herpesvirus type 4, feline herpesvirus, canine herpesvirus, and herpes simiae virus. Equine herpesviruses 1 and 3, human herpesviruses 1 and 2, and herpes simiae virus were all sensitive to BIOLF-62 at concentrations of less than 0.55 micrograms/ml. Equine herpesvirus types 1 and 3 were particularly sensitive, viral median effective dose (ED50) concentrations of the drug being only 0.033 and 0.16 micrograms/ml, respectively. Such high antiviral potency and low cell toxicity indicate that BIOLF-62 might be useful in the treatment of infected animals.

Acyclovir↗
PubMed1982

A new nucleoside analog, 9-[[2-hydroxy-1-(hydroxymethyl)ethoxyl]methyl]guanine, highly active in vitro against herpes simplex virus types 1 and 2.

A novel nucleoside analog, 9-[[2-hydroxy-1-(hydroxymethyl)ethoxy]methyl]-guanine (BIOLF-62), was found to have potent antiviral activity against herpes simplex virus types 1 and 2 at concentrations well below cytotoxic levels. For example, the Patton strain of herpes simplex virus type 1 was susceptible at concentrations 140- to 2,900-fold below that which inhibited cell division by 50%, depending upon the cell line used for assay. Different herpesvirus strains varied considerably in their susceptibility to the drug, as did results obtained with the same virus strain in different cell lines. BIOLF-62 compared favorably with 5-iodo-2'-deoxyuridine and acyclovir with respect to ratios of viral to cell inhibitory drug concentrations. Patterns of drug resistance to herpesvirus mutants suggested that the primary mode of action of BIOLF-62 is different from that of known antiviral compounds. Human adenovirus type 2, varicella-zoster virus, and Epstein-Barr virus were inhibited by this drug but at concentrations within the cell inhibitory range. Vaccinia virus and human cytomegalovirus were not inhibited at high drug concentrations.

Antiviral Agents↗
PubMed1982

Synergism among BIOLF-62, phosphonoformate, and other antiherpetic compounds.

9-[[2-Hydroxy-1-(hydroxymethyl)ethoxy]methyl]guanine (BIOLF-62) is highly synergistic with either phosphonoformate or phosphonoacetate when used in combination against herpes simplex virus types 1 and 2 in vitro. Acycloguanosine did not show significant synergism with these two compounds. Bromovinyldeoxyuridine and phosphonoformate were highly synergistic against herpes simplex virus type 2, but not against type 1.

Acyclovir↗
PubMed1981

Visualization of minute centers of viral infection in unfixed cell cultures by an enzyme-linked antibody assay.

Enzyme-linked antibody was used to treat unfixed herpesvirus-infected human fetal lung cell cultures in a mode which permitted the visualizing of local sites of infection. Foci containing as few as 20 herpesvirus-infected cells produced sufficient viral mass to be easily detectable by this method. 'Clouds' or 'plumes' of colored reaction product diffused into the substrate overlay, accumulated above and around each focus of infection and allowed quantitation of the number of foci in a culture. The number of minute centers of viral infection determined by the enzyme-linked antibody method corresponded almost exactly with values obtained by fluorescence microscopy. Quantitation of herpes simplex infectivity by focus assay was possible within only 17 h after culture inoculation, well before cytopathic effects were visible macroscopically. The technique was also applied to demonstrate measles and mumpsvirus plaques (infectious centers) in Vero cell cultures.

Animals↗
PubMed1981

Rapid assay of herpes simplex virus sensitivity to drugs.

A 16 h immunofluorescence microplaque reduction assay for herpes simplex virus (HSV) was compared with a plaque reduction assay to assess its feasibility as a rapid technique for determining viral sensitivity to drugs. A drug-sensitive HSV strain (HSV-1 Patton) and a drug resistant strain (HSV-1-P-ACG-R), both grown in cell cultures, were compared against serial dilutions of adenine arabinoside (Ara-A) and acycloguanosine (ACG). Similar studies were done with virus obtained directly from experimentally infected rabbit corneas. The microplaque assay yielded results similar to the plaque assay and has the advantage of speed, which may be useful in clinical situations.

Acyclovir↗
PubMed1981

Differentiation of members of the human herpesviridae family by radioimmunoassay.

Many individuals who are seronegative for one member of the human Herpesviridae family are strongly seropositive for other members. Using sera from such individuals, the radioimmunoassay technique demonstrated absence of antigen-antibody cross-reactions between varicella-zoster virus (VZV) and herpes simplex virus (HSV) at levels of less than one part in 1,000. Sera containing antibody to both HSV and VZV were absorbed with antigens of one agent without significantly altering the amount of remaining antibody to the other antigen. This further suggests that HSV and VZV do not share a common antigen. The same radioimmunoassay technique and serum absorption method that revealed no serological cross-reactions between HSV and VZV revealed the expected cross-reaction between HSV type 1 (HSV-1) and HSV-2. Reciprocally absorbed anti-HSV-1 and anti-HSV 2 sera were able to differentiate the two HSV types reliably. No cross-reactions were seen between cytomegalovirus, VZV, and HSV or between Epstein-Barr virus, VZV, and HSV. We postulate that heterotypic antibody responses sometimes observed for VZV after primary infections by HSV may not be due to shared antigens, but to activation of latent VZV infections, release of new VZV antigens, and consequent stimulation of new antibody production to VZV.

Antigens, Viral↗
PubMed1981

Cryopreservation of varicella-zoster virions without loss of structural integrity or infectivity.

Varicella-zosterer virions present in infected cells or in a cell-free state were freeze-dried without loss of structural integrity of infectivity. Generally, yields of greater than 5 log10 foci/ml (infected cells) or greater than 4 log10 PFU/ml (cell-free virus) were recovered from varicella-zoster virus-infected human melanoma cells both before and after lyophilization in phosphate-buffered media containing 0.1-1.0 M sucrose. Virus frozen in solutions lacking sugar had little or no residual infectivity after vacuum sublimation was completed. Visualization by electron microscopy demonstrated large numbers of enveloped virions in the virus preparations lyophilized in media containing sucrose; in marked contrast, virus subjected to freeze-drying in buffered solutions without sugar consisted mainly of naked nucleocapsids. Water analyses by Karl Fischer titration suggested that residual moisture retained by sugar prevented disenvelopment of the varicella-zost virion.

Edetic Acid↗
PubMed1980

In vitro and in vivo resistance of herpes simplex virus to 9-(2-hydroxyethoxymethyl)guanine (acycloguanosine).

In vitro passage of the Patton strain of herpes simplex virus type 1 (HSV-1-P) in the presence of acycloguanosine (ACG) led to the emergence of a highly drug-resistant strain (HSV-1-P-ACG-R). Over 1% of virions in 6 of 15 HSV strains tested were able to form plaques in the presence of 10 mumol of ACG on first exposure to the drug. Therefore, there exists among HSV strains a broad range of susceptibilities to ACG, and some strains contain particles which are partially resistant to ACG before ever contacting the drug. HSV-1-P-ACG-R was partially resistant to iododeoxyuridine; conversely, iododeoxyuridine-resistant virus was highly resistant to ACG. ACG-resistant virus (HSV-1-P-ACG-R) was equal to its parent strain (HSV-1-P) in susceptibility to adenine arabinoside. The HSV-1-P-ACG-R strain produced corneal lesions in rabbits which were completely refractory to topical treatment with 1% unguent ACG, but responsive to 3% ACG. Partially resistant HSV could be isolated from the eyes of rabbits infected with ACG-susceptible virus and treated topically with either 1 or 3% ACG for 6 days.

Acyclovir↗
PubMed1980

Serum antibodies to herpes simplex virus type 1 during active oral herpes infection.

Subjects with oral herpes lesions at the time of serum sampling had higher-efficiency antibody (higher proportion of neutralizing antibody as determined by plaque reduction, compared with total antibody as detected by radioimmunoassay) to herpes simplex virus type 1 (HSV-1) than did subjects with no lesions at the time of serum sampling. These higher-efficiency sera also had higher antibody titers to structural components of herpes simplex virus type 1 than did the low-efficiency sera. Absorption of high- and low-efficiency sera with purified herpes simplex virus type 1 particles removed all neutralizing antibody but not all antibody detected by radioimmunoassay. High-efficiency serum was depleted of more antibody to particulate antigen that was the low-efficiency serum, indicating that the high-efficiency serum contained a higher proportion of antibody to the virus particle.

Adult↗
PubMed1980

A serological study of herpes simplex virus type 1 antibody over a 13-year period.

HSV-1 serum antibody titers determined by 50% plaque reduction and confirmed by radioimmunoassay in 11 samples from an individual over a 13-year period indicated a significant increase between the first sample and a sample taken 9 years later. This increase did not seem to reflect loss of antibody in the low titered serum sample due to storage.

Antibodies, Viral↗
PubMed1980

Rubella immunity: comparison of hemagglutination inhibition and radioimmunoassay antibody methods.

A threshold (1:10) rubella antibody hemagglutination inhibition (HAI) titer was obtained for 288 of 6537 (4.4%) obstetric patients. Random sera from 84 of these patients were compared for rubella antibody by both HAI and a very sensitive radioimmunoassay (RIA) technique. By RIA, 17% of the sera had no detectable rubella antibody, suggesting that those patients were truly susceptible to rubella. In addition, 8 of 55 paired prevaccine and postvaccine sera with titer increases demonstrated by HAI were analyzed by RIA. Seven of the 8 prevaccine sera were shown to have no rubella virus antibody by RIA, and RIA showed seroconversion in all 8 postvaccine sera.

Antibodies, Viral↗