Biomedical subjects
Publications and source records attributed to J Dennis.
"X Lite", a thermoplastic material was used in eight patients as a splint for protecting split skin grafts on the back. After an average stay of 4.3 days patients were discharged with no incidence of graft failure. The patients involved felt comfortable whilst remaining supine. We believe time saved by adequate protection leading to successful grafts makes this method economically useful.
Molluscum contagiosum virus (MCV) is an unclassified poxvirus which has recently become recognized as causing a major sexually transmitted disease. At present no assay is available for specific detection of MCV because the virus cannot be serially propagated in cell culture. Since MCV produces an abortive, limited growth with some cytopathic effect in certain cell lines, we were able to develop an in situ hybridization assay for detection of MCV genome in clinical specimens. Human fetal diploid lung cell monolayers were infected with clinical specimens, and after proper incubation and fixation in paraformaldehyde, hybridization was performed under full stringency conditions with a molecularly cloned biotinylated probe. Only MCV infected cells showed homology to the MCV probe with a purple-brown cytoplasmic staining. Additionally, we have described an in situ hybridization assay for direct detection of MCV genome in formalin-fixed, paraffin-embedded biopsies. Characteristic intracytoplasmic Molluscum bodies (Henderson-Paterson bodies) were detected in stratum spinosum cells of the epidermis. Striking staining similarities have been observed between in situ hybridization and haematoxylin-eosin cytostaining. These procedures are the first successful identification of MCV genome in clinical samples by molecular hybridization, with sensitivity and specificity equal to or greater than electron microscopy.
Periconceptional supplementation with Pregnavite Forte F was offered to women who presented consecutively to the Oxford genetic counselling service in the early 1980s who had previously had one or more pregnancies complicated by a neural tube defect. The first 100 children born alive to these women are the subject of this study. Birth weight, gestation, and congenital abnormalities were recorded. At age 2-5 years all 96 children remaining in the United Kingdom were assessed clinically and developmentally and behavioural information was obtained by questionnaire. At age 7-10 years, follow up of 91 children by telephone and postal questionnaire yielded further information about growth, general health, vision, hearing, and educational and behavioural status. Entry criteria excluded single mothers but the social class distribution of the sample was otherwise representative of the Oxfordshire population. There were no recurrences of neural tube defects. One child had radiological evidence of spina bifida occulta affecting only the fifth lumbar vertebra. One had an autosomal recessive disorder. Eight had random minor congenital anomalies. Birth weight for gestational age was significantly greater than for the local population and at age 7-10 years the girls were considerably taller than expected. Health, auditory, visual, and developmental status were no different from the general population. None of the children had special educational needs. None showed a major behaviour disorder but worries, fussiness, and fearfulness were highly significantly over represented.
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Three fractions of poly(A)-containing RNA were separated from total rat liver RNA using poly(U)-Sepharose 4B affinity chromatography. The poly(A)-containing RNA fractions were released by thermal elution. Fraction 1, eluted under the mildest conditions, and had poly(A) tracts of approx. 200 AMP units in length which appeared to be associated with poly(U) sequences of 20-50 UMP in length. Fraction 1 appeared to be present mainly in the nucleus and, its size distribution was similar to that of fractions 2 and 3. Fractions 2 and 3 eluted at higher temperatures and were associated mainly with polysomal and microsomal fractions. Poly(U) sequences were absent in fractions 2 and 3 while their poly(A) sequences had a size distribution characteristic of those reported in the mRNA of other organisms.
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To explore possible advantages which immunoperoxidase (IP) staining might have over immunofluorescence (IF) staining for identifying rubella virus isolates, direct comparative studies were done on the same coded clinical materials using the same rubella immune rabbit serum as the primary antiserum in both systems. The rubella immune rabbit serum and conjugated anti-rabbit immune globulins could be used more dilute in the IP system than in the IF system. Both IP and IF staining detected rubella antigen in all specimens which were positive by interference. IP staining also detected low levels of rubella antigen in a few additional specimens which had originally been positive for rubella virus, but which on retesting were negative by interference and IF staining. With second-cell-culture-passage material, IP and IF staining showed comparable specificity, and the few specimens which reacted nonspecifically generally did so in both systems. Cell cultures inoculated directly with urine specimens showed greater nonspecificity by IP than by IF, but this activity could be abolished by pretreatment with sodium azide and peroxide; other methods tried for inactivating endogenous peroxidase activity destroyed rubella antigen as well. The intensity of staining for positive specimens was comparable in the two systems. However, more antigen was demonstrable in both systems when BHK-21 cells were inoculated as a cell suspension and then permitted to grow into monolayers than when the same specimens were inoculated into preformed monolayers. IP staining was considered to be a highly satisfactory alternative to IF staining for identification of rubella virus isolates.
An enzyme immunossay (EIA) was adapted for detection of antibody to varicella-zoster virus, and its sensitivity and specificity were compared with those of neutralization, immune adherence hemagglutination (IAHA), and complement fixation tests. Test sera showed little nonspecific reactivity in the EIA system, and valid results could usually be obtained at serum dilutions as low as 1:8. Demonstration of the presence or absence of varicella-zoster viral antibody by EIA showed 94% correlation with results obtained in neutralization tests, but EIA titers were 2- to 16-fold higher than neutralizing antibody titers. Results by IAHA showed 87% correlation with those obtained by neutralization. No false positive IAHA results were seen, but a number of false negative IAHA results were seen at the 1:8 serum dilution, particularly in older individuals. With increasing age (>40 years), and presumably increased time from varicella infection, neutralizing antibody levels generally declined to 1:8 or 1:16, EIA levels fell to 1:128 or 1:256, and IAHA and complement fixation antibody titers were usually <1:8 or 1:8. EIA and IAHA were as reliable as the neutralization and complement fixation tests for serodiagnosis of varicella and zoster infections. All tests demonstrated heterotypic varicella-zoster antibody titer rises in selected patients with initial herpes simplex virus infections, but fewer heterotypic responses were seen by EIA than by the other methods. EIA offers a rapid, sensitive, and specific method for varicella-zoster antibody assay that is applicable to use in a clinical setting.
Various subunit antigens of varicella-zoster (V-Z) virus were examined for complement-fixing (CF) activity with sera from homotypic infections and from herpes simplex virus (HSV) infections in which a CF antibody titer rise was demonstrated with crude V-Z antigen. The subunit antigens included nucleocapsids, envelopes, a soluble antigen produced from infected culture fluids by sucrose density gradient centrifugation, a soluble antigen produced by reducing the volume of clarified infected culture fluids, a soluble antigen derived from infected cell lysates, a "viral" antigen consisting largely of enveloped particles with a few nucleocapsids, and a cell membrane-associated antigen. None was more suitable than crude V-Z antigen for serodifferentiation of V-Z virus and HSV infections. The envelope antigen, cell membrane antigen, and the soluble antigen prepared by density gradient centrifugation showed little reactivity with sera from varicella and HSV infections, but gave high antibody titers with sera from zoster infections, suggesting that a secondary V-Z virus infection is required to produce an antibody response to these subunit antigens. Patients with varicella and zoster infections and the selected patients with HSV infections all showed significant CF antibody responses to the other V-Z subunit antigens.
To our knowledge, there has been reported only one previous case of prosthetic valve endocarditis due to Neisseria meningitidis. In describing a second fatal case, we emphasize the difficulty of clinical diagnosis due to lack of characteristic high fever, murmur, skin rash, or splenomegaly. This lack of findings plus the chronic, intermittent course suggest a rare occurrence of endocarditis complicating chronic meningococcemia with an acute exacerbation, leading to rapid decompensation and death.
Foci of cells infected with human cytomegalovirus were noted to stain more intensely than uninfected cells with neutral red, and this provided the basis for development of a plaque assay and plaque reduction neutralization test for cytomegalovirus. Plaques demonstrable by neutral red staining could be counted at 8 days after infection; thus, results could be obtained earlier than for plaque assay systems based upon the viral cytopathic effect, a fewer manipulations were required for staining cell monolayers to demonstrate plaques. Certain variables affecting plaque size and numbers and antibody titers were defined. Addition of fresh guinea pig complement to the reaction mixtures markedly enhanced cytomegalovirus-neutralizing antibody titers of hyperimmune animal sera, but titers of human sera were enhanced only two-or fourfold.
Expired ventilation (VE), tidal volume (VT), frequency (f), and alveolar PCO2 (PACO2) were examined in six normal infants at 41 to 52 weeks post-conceptional age and in two infants who were apneic at birth. Their response to breathing 5% carbon dioxide in air and to 100% oxygen in quiet sleep were compared to those in rapid eye movement (REM) sleep. VE in normal infants was 259 ml/kg/min in REM and 200.2 ml/kg/min in quiet sleep with the difference being due to decreased carbon dioxide production and to decreased dead space. VE increased 34.4 ml/kg/min/mm Hg of PCO2 elevation with 5% carbon dioxide breathing during REM and was not significantly different during quiet sleep. During oxygen breathing VE fell by 32.7% at 30 seconds before increasing again. In the affected infants, VE and PACO2 during REM at 1 and 4 months were normal. At 1 month, during quiet sleep, each infant became apneic and PACO2 rose 9 and 8 mm Hg/min respectively. At this time mechanical ventilation was begun. At 4 months, during quiet sleep, VE was 0.064 and 0.063 ml/kg/min at PACO2 of 66 mm Hg in each infant. The change was due entirely to a decrease in VT to 2.3 and 2.5 ml/kg. At this time 5% carbon dioxide breathing given during normal ventilation in REM produced an abrupt fall in VT to 2.0 and 2.2 ml/kg with no change in frequency. Oxygen breathing during REM at one month had no effect but at 4 months produced apnea requiring mechanical ventilation after one minute. The findings suggest that the ventilatory response to carbon dioxide is (1) important in initiation of extrauterine ventilation and (2) in sustaining ventilation particularly in quiet sleep. It is not necessary in sustaining ventilation awake or in REM sleep and it represents a balance between the stimulatory and depressant effects of carbon dioxide on the central nervous system.
Simple, inexpensive plastic templates have been devised to aid in proper identification of cell cultures in microtiter plates during inoculation and microscopic observation. These have resulted in greater accuracy in performing and reading tests and in a saving of the microbiologists' time.
A multi-purpose, inexpensive laboratory light box is described which can be used for observing and photographing results of a diversity of laboratory tests using either white or ultraviolet light.