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Syngnathidae (seahorses, seadragons and pipefish) suffer significant losses from non-tuberculous mycobacteria. However, they produce markedly different lesions in response to the disease compared to other teleost species, notably infrequent granuloma formation. This study evaluated 270 syngnathid fish, from which 92 were diagnosed with mycobacteriosis by histopathology, culture or both. Microscopic lesions variably consisted of random foci of coagulative necrosis in multiple organs, containing high numbers of free bacteria and large aggregates or sheets of macrophages with cytoplasm laden with acid-fast bacilli. Mycobacterial associated granulomas were identified in only six seahorses. Five fish had positive cultures with no observed microscopic changes. RNA-seq of the head kidney was performed to investigate the transcriptome of two infected and six non-infected lined seahorses Hippocampus erectus. Assembled and annotated putative transcripts serve to enrich the database for this species, as well as provide baseline data for understanding the pathogenesis of mycobacteriosis in seahorses. Putative components of the innate immune system (IL-1β, IL-6, TNF, NOS, Toll-like receptor 1, MHC Class I, NF-κβ, transforming growth factor beta, MyD88) were identified in the RNA-seq data set. However, a homolog for a key component in the TH1 adaptive immune response, interferon-gamma, was not identified and may underlie the unique pathologic presentation., (© 2017 John Wiley & Sons Ltd.)

A number treatment options are emerging for patients with retinal degenerative disease, including gene therapy, trophic factor therapy, visual cycle inhibitors (e.g., for patients with Stargardt disease and allied conditions), and cell transplantation. A radically different approach, which will augment but not replace these options, is termed neural prosthetics ("artificial vision"). Although rewiring of inner retinal circuits and inner retinal neuronal degeneration occur in association with photoreceptor degeneration in retinitis pigmentosa (RP), it is possible to create visually useful percepts by stimulating retinal ganglion cells electrically. This fact has lead to the development of techniques to induce photosensitivity in cells that are not light sensitive normally as well as to the development of the bionic retina. Advances in artificial vision continue at a robust pace. These advances are based on the use of molecular engineering and nanotechnology to render cells light-sensitive, to target ion channels to the appropriate cell type (e.g., bipolar cell) and/or cell region (e.g., dendritic tree vs. soma), and on sophisticated image processing algorithms that take advantage of our knowledge of signal processing in the retina. Combined with advances in gene therapy, pathway-based therapy, and cell-based therapy, "artificial vision" technologies create a powerful armamentarium with which ophthalmologists will be able to treat blindness in patients who have a variety of degenerative retinal diseases.

Immunohistochemistry of tumour samples is increasingly used in the triage of families where hereditary non-polyposis colorectal cancer (HNPCC) due to mismatch repair defects is suspected. Usually, this is undertaken in tumours that are a recognised part of the spectrum of HNPCC-related cancers e.g. colon or endometrial cancers. Although breast cancers are not classed as part of this spectrum, this study examined the extent to which some breast tumours do arise by the mismatch repair pathway in these families. This may have clinical utility in families where an individual with a 'classic HNPPC-related' tumour is not available for evaluation. Immunohistochemistry of a breast tumour may identify an individual in whom germline mutation testing is worthwhile.

The selection of candidates for BRCA germline mutation testing is an important clinical issue yet it remains a significant challenge. A number of risk prediction models have been developed to assist in pretest counselling. We have evaluated the performance and the inter-rater reliability of four of these models (BRCAPRO, Manchester, Penn and the Myriad-Frank). The four risk assessment models were applied to 380 pedigrees of families who had undergone BRCA1/2 mutation analysis. Sensitivity, specificity, positive and negative predictive values, likelihood ratios and area under the receiver operator characteristic (ROC) curve were calculated for each model. Using a greater than 10% probability threshold, the likelihood that a BRCA test result was positive in a mutation carrier compared to the likelihood that the same result would be expected in an individual without a BRCA mutation was 2.10 (95% confidence interval (CI) 1.66-2.67) for Penn, 1.74 (95% CI 1.48-2.04) for Myriad, 1.35 (95% CI 1.19-1.53) for Manchester and 1.68 (95% CI 1.39-2.03) for BRCAPRO. Application of these models, therefore, did not rule in BRCA mutation carrier status. Similar trends were observed for separate BRCA1/2 performance measures except BRCA2 assessment in the Penn model where the positive likelihood ratio was 5.93. The area under the ROC curve for each model was close to 0.75. In conclusion, the four models had very little impact on the pre-test probability of disease; there were significant clinical barriers to using some models and risk estimates varied between experts. Use of models for predicting BRCA mutation status is not currently justified for populations such as that evaluated in the current study.

Asusceptibility testing program was established to determine the prevalence of resistance to penciclovir among herpes simplex virus isolates collected from patients participating in 11 world-wide clinical trials involving penciclovir (topical or intravenous formulations) or famciclovir, the oral prodrug of penciclovir. These trials represented nine randomised double blind, placebo or aciclovir-controlled studies and two open-label studies. Groups surveyed included immunocompetent or immunocompromised patients receiving 2 to 12 months chronic suppressive therapy for genital herpes, immunocompetent patients with recurrent herpes labialis treated for four days, and immunocompromised patients with mucocutaneous herpes simplex virus (HSV). Another subset of patients had been identified as non-responders to aciclovir or to valaciclovir. This program assessed the susceptibility profile for a total of 2145 herpes simplex virus isolates from 913 immunocompetent and 288 immunocompromised patients treated with penciclovir, famciclovir, aciclovir or placebo (depending on trial design). HSV isolates were tested for susceptibility to penciclovir using the plaque reduction assay (PRA) in MRC-5 cells. Resistance was defined as an IC(50)>or=2.0 microg/ml or an IC(50)> 10-fold above the wild type control virus IC(50) within that particular assay. Penciclovir-resistant HSV was isolated from 0.22% immunocompetent patients, and 2.1% of immunocompromised patients overall and therefore the frequency of penciclovir-resistant herpes simplex virus in the immunocompetent population approximates that of aciclovir-resistant herpesvirus reported previously. Penciclovir-resistant HSV isolates were more common in isolates from immunocompromised patients, consistent with aciclovir clinical experience. Treatment with penciclovir (intravenous formulation) was associated with the development of resistant HSV in only one severely immunocompromised patient (day 7 isolate IC(50) = 2.01 microg/ml), although treatment was effective and resulted in the complete clearance of the lesion by day 8. No patients receiving topical penciclovir developed treatment-associated penciclovir-resistant HSV, and a single immunocompromised patient developed resistant HSV upon treatment with oral famiciclovir.

Helicobacter pylori infection is linked to chronic gastritis, peptic ulcer and gastric carcinoma. During H. pylori infection, class II MHC expression by the gastric epithelium increases, as does the number of local CD4(+) T cells, which appear to be important in the associated pathogenesis. These observations suggested that the epithelium might present antigens to T cells. Thus, we sought to determine whether gastric epithelial cells process antigens to establish their function as local antigen presenting cells (APC). We examined a panel of gastric epithelial cell lines for expression of the antigen processing cathepsins B (CB), L (CL), S (CS), and D (CD). The mRNA for these enzymes were detected by RT-PCR and the enzymes in the gastric epithelial cells were identified by various independent methods. We corroborated the expression of CB and CD on gastric epithelial cells from human biopsy samples. The functions of these proteases were confirmed by assessing their ability to digest ovalbumin, a conventional dietary antigen, and proteins from H. pylori. In summary, multiple lines of evidence suggest gastric epithelial cells process antigens for presentation to CD4(+) T cells. To our knowledge, these are the first studies to document the antigen processing capacity of human gastric epithelial cells.

Objective: Streptococcus pneumoniae is one of the most clinically significant pathogens with emerging antibiotic resistance. We performed a surveillance study in isolated rural populations of healthy children to estimate the prevalence of pneumococcal resistance and to contrast factors that predict pneumococcal carriage with those that specifically predict resistant pneumococcal carriage., Methods: The study was conducted in 1998 in 2 rural communities in Utah. Families were recruited directly for participation through community canvassing. Surveillance nasopharyngeal cultures were obtained from children who were younger than 8 years. Antibiotic usage and information on other potential risk factors were obtained from questionnaires and local pharmacy records. Resistance was determined by testing isolates for susceptibility to penicillin, cefaclor, trimethoprim-sulfamethoxazole, erythromycin, ceftriaxone, and trovafloxacin. Selected resistant isolates were characterized further by serotyping, pulsed field gel electrophoresis, and Southern blot with DNA probes specific for the pneumococcal lytA gene and for antibiotic resistance genes., Results: In April 1998, surveillance nasopharyngeal cultures were obtained from 368 children aged

Progressive genetic changes such as the inactivation of tumor suppressor genes (TSG) are thought to play an important role in the initiation and progression of ovarian cancer. Frequent nonrandom allelic imbalance (AI) at 8p11-p21 and 8p22-pter suggests the existence of TSGs that may be involved in the carcinogenesis of several human malignancies. We investigated 70 ovarian tumors with 11 highly polymorphic markers spanning 8p12-p21 and 8p22-pter to produce an AI map of 8p in epithelial ovarian cancer. Allelic imbalance was demonstrated in 54 tumors (77%), most frequently occurring at D8S136 (54%) and at D8S1992 (55%). Poorly differentiated and advanced stage cancers were more often affected by AI (G1+G2 vs. G3; 20% vs. 66%; stage I+II vs. III+IV, 36% vs. 54%, P<.001; Kruskal-Wallis test) than well differentiated and early stage tumors. There was no relationship between histological subtype and AI. Smallest regions of overlap (SRO) were delineated by analyzing 38 tumors with partial AI. This study provides compelling evidence for the involvement of TSGs on the short arm of chromosome 8, at 8p12-p21 and at 8p23 in the development and progression of epithelial ovarian cancer.

The identification of mutations in the BRCA1 gene poses difficulties in achieving a screening outcome that satisfies the twin needs of speed and accuracy. These needs must also take into account the patient's family history and the statistical evaluation of the probability of detecting a mutation. Given the above, we present here a hierarchical mutation screening strategy that comprises two tiers: first, multiplex heteroduplex and exon 13 duplication analysis; second, exon amplification and direct sequencing using a 96-well tray format. The advantages of this strategy are two-fold: first, the division of analytical tools in order to achieve low and high-resolution mutation screening, respectively; second, a streamlined sequencing approach that leads to a sensitive and rapid assay that reduces labor costs and handling errors. The success of this approach is shown by the identification of a novel deletion mutation in exon 14 of the BRCA1 gene, which was not detected by the more conventional protein truncation assay due to the small size of the predicted truncated protein., (Copyright 2000 Wiley-Liss, Inc.)

Short-rib polydactyly dysplasia (SRP) is an autosomal recessive, lethal skeletal dysplasia. Sonographic assessment of subsequent pregnancies is, therefore, recommended. This case indicates that this diagnosis can be made in the latter part of the first trimester. A 30-year-old multigravid woman presented at 13 weeks' gestation for an ultrasound examination. She had had a termination of pregnancy for a fetus with pathologically confirmed short-rib polydactyly dysplasia, type I (Saldino-Noonan). On transvaginal sonography, a narrow chest, symmetrical micromelia, polydactyly and anasarca were present. An autopsy confirmed recurrent SRP. Short-rib polydactyly dysplasia may be diagnosed with transvaginal sonography in the first trimester.

The Claybury Nursing Stress Study assessed the levels of occupational stress experienced by both ward-based and community nurses. This paper presents the results obtained from a qualitative analysis of statements made by community psychiatric nurses during a questionnaire based interview. Key areas identified by CPNs as relevant to stress and coping are reported and suggestions for further, qualitative research in this area are made.

A randomised trial of simvastatin and enalapril in patients with chronic renal failure on dialysis: effects on serum lipoprotein concentrations and left ventricular mass. Left ventricular hypertrophy and abnormalities of lipoprotein metabolism are both possible contributors to the high risk of cardiovascular death in patients with chronic renal failure on dialysis. We investigated the effects of simvastatin on lipid and lipoprotein concentrations and the effects of enalapril on left ventricular mass in 107 patients receiving haemodialysis or continuous ambulatory peritoneal dialysis. Patients were randomised in a factorial design to receive simvastatin (10 mg daily) or placebo and enalapril (2.5-5 mg daily) or placebo. During follow-up, there was a significant excess of patients withdrawn from enalapril because of hypotension (2p = 0.002), and after 6 months only 55% of those assigned enalapril were still on treatment. From baseline to 6 months, there were no statistically significant differences in left ventricular mass or left ventricular dimensions between patients assigned enalapril and those assigned placebo. Among the patients assigned simvastatin, total cholesterol was reduced by 13% (2p = 0.001), LDL cholesterol was reduced by 17% (2p = 0.003) and apolipoprotein B was reduced by 12% (2p = 0.005) compared to patients assigned placebo. There were borderline significant (2p = 0.05 to 0.08) reductions in VLDL cholesterol, total triglyceride and VLDL triglycerides of 26%, 12% and 17% respectively. Large-scale trials are now required to determine whether reductions in lipid and lipoprotein concentrations confer a reduction in coronary heart disease morbidity and mortality in patients on dialysis.

Objective: To determine the frequency with which one or both normal ovaries can be visualized during a routine obstetric ultrasound examination., Methods: The population consisted of 5617 pregnant women at 5.0-39.9 weeks' gestation, studied cross-sectionally. The sonographic visualization rate for one or both normal ovaries, as well as their position above or below the level of the umbilicus, was recorded for one examination in each patient., Results: The study population was divided into three groups according to gestational age: first trimester, 5.0-12.9 weeks; second trimester, 13.0-26.9 weeks; and third trimester, 27.0-39.9 weeks. There were 829, 3195, and 1593 women in the first, second, and third trimesters, respectively. Most women were examined transvaginally in the first trimester; transabdominal sonography was used in the second and third trimesters. The ability to visualize one or both ovaries declined significantly (P < .05) from the first trimester to the second, as well as from the second trimester to the third (P < .001). The percentage of ovaries that were visualized above the umbilicus increased from 2.4% in the first trimester to 10.1% in the second trimester (P < .001), and to 21.7% in the third trimester (P < .001)., Conclusion: As gestational age advances, there is a significant reduction in the ultrasound visualization rate of normal ovaries. This investigation provides normative data for ovarian visualization throughout pregnancy that may be helpful in establishing ultrasound laboratory standards.

In this paper we present data from three research studies on stress, coping and burnout in mental health nurses. All three studies used a range of self report questionnaires. Measures included a demographic checklist, the General Health Questionnaire (GHQ-28), the Maslach Burnout Inventory, the DCL Stress Scale and the Cooper Coping Skills Scale. In all, 648 ward based mental health nurses were surveyed. There were no significant differences between levels of psychological distress on GHQ Total Score, but there were differences in caseness rates. In Study 3, some 38% of nurses were found to score at or above the criterion for caseness. The main stressors for ward staff were to do with staff shortages, health service changes, poor morale and not being notified of changes before they occurred. Differences in coping skills were found across studies. The study group with the highest stress scores also had the lowest coping skills scores. This was also associated with significantly higher alcohol consumption and greater self reported sickness absence. Scores on the Maslach Burnout Inventory showed higher levels of burnout amongst nurses in Study 3. These three studies have confirmed that stress is a problem for ward based mental health nurses. Two main implications arise from this work. Firstly we need models of the stress process that are empirically based, and which help us identify the moderating variables that reduce the impact of stressors on nurses. Secondly, we need to utilise this knowledge to deliver stress management interventions for staff. We end by outlining a model which may help us both understand the process of stress causation, and move towards our goal of stress reduction.

The Claybury community psychiatric nurse (CPN) stress study collected data on stress levels in 250 CPNs and 323 ward-based psychiatric nurses (WBPN) in the North East Thames region. Four out of 10 CPNs were found to be experiencing high levels of psychological distress on GHQ scores. Whilst both CPNs and WBPNs scored highly on scores of occupational burnout, especially on emotional exhaustion scores, WBPNs scored worse on emotional detachment from their patients and were achieving less personal fulfilment from their work. Both groups of nurses were more satisfied with direct patient clinical work than with their employment conditions, particularly their working environments and, for CPNs, their relationships with their managers. The different patterns of coping skills are explored and discussed for both groups of nurses, especially the use of social support, time management and organization of tasks. The study concludes that whilst major changes are occurring in the psychiatric arena for both groups of nurses, stress is reaping its toll on mental health nurses, in terms of higher absence rates, lower self-esteem and personal unfulfilment. This could not only affect the quality of patient care but also future career prospects for nurses. The study invites serious consideration of introducing stress-reducing measures in the work-place as well as further research into specific stressors for different groups of nurses.

Work place stress is an increasing concern for mental health nurses. This article reports the findings of two research studies of ward and community-based staff, identifying specific stressors. Reducing staff stress levels is essential if patient care is not to suffer.

With the development of the concept of community care there has been a significant expansion of the community psychiatric nurse (CPN) profession. The present study attempts to examine which aspects of their work CPNs currently find stressful. The study also examines the various strategies which CPNs feel to be useful in attempting to cope with such occupational stress. Forty-four CPNs in four health districts participated in this Q-methodological study which provided the opportunity for CPNs to construct their own concepts of stressors and coping strategies. The results obtained indicated that CPNs identified nine distinct areas of stress within their work, along with 12 distinct coping strategies which they considered useful in attempting to deal with such stress. The implications of these findings are discussed.

The issue of stress amongst health care professionals is currently a major concern within the British National Health Service. It is important for researchers to develop both reliable and valid psychometric measures to assess occupational stress. This paper outlines the development and piloting of a measure of professional stress for community mental health nurses, the CPN Stress Questionnaire (Revised). Data on the reliability and validity of this new measure are presented. It is concluded that this measure has good psychometric properties. A range of possible research applications is outlined.

Rat ovarian surface epithelial cells transformed spontaneously in vitro have been found to have homozygous deletions of the interferon alpha (IFNA) gene. This suggests that inactivation of a tumor-suppressor gene in this region may be crucial for the development of ovarian cancer. We therefore used microsatellite markers and Southern analysis to examine the homologous region in humans--the short arm of chromosome 9--for deletions in sporadic ovarian adenocarcinomas and ovarian tumor cell lines. Loss of heterozygosity occurred in 34 (37%) of 91 informative sporadic tumors, including some benign, low-malignant-potential and early-stage tumors, suggesting that it is an early event in the development of ovarian adenocarcinoma. Furthermore, homozygous deletions on 9p were found in 2 of 10 independent cell lines. Deletion mapping of the tumors and lines indicates that the candidate suppressor gene inactivated as a consequence lies between D9S171 and the IFNA locus, a region that is also deleted in several other tumors and that contains the melanoma predisposition gene, MLM.

Inactivation of the DCC gene on chromosome 18 owing to loss of heterozygosity is a common finding in colorectal cancer. Because both ovarian and colon cancer are features of Lynch syndrome II, which has been provisionally mapped to chromosome 18, we hypothesized that loss of heterozygosity at the DCC locus may also occur in ovarian neoplasia. Fifty-two sporadic ovarian adenocarcinoma tumours were analysed by Southern blotting for loss of heterozygosity (LOH) at six chromosome 18 loci. Overall, tumours from 31 patients (60%) showed allelic loss at one or more of these loci. A similarly high level of LOH, 66%, was found at D17S5 (17p13.3). In contrast, moderate levels of LOH, of 31%, 39% and 33%, were found at MYCL1 (1p32), D1S57 (1p) and D14S20 (14q32.33) respectively. However, analysis of partial chromosome deletions in 11 patients indicates that the smallest region of overlap appears to exclude the DCC gene but to be between the D18S5 and D18S11 loci. This suggests that another locus, as well as or apart from DCC, may be involved.

Monoclonal antibodies were generated against idiotopes on an NK target antigen-specific IgM monoclonal antibody (mab). This mab (18C2) was originally produced against (NC-37) human EBV-transformed B cells. The 18C2 mab inhibits natural killer cell lysis of NC-37 and other target cells by preventing conjugate formation. Anti-18C2(id) mabs were tested for binding to effector cells and screened by ELISA, flow cytometry, and by inhibition of NK cytotoxicity. Two of the anti-18C2(id) (anti-id) mabs (12H1.C5 and 6D9.B11) were chosen for further study. The idiotypic specificity of these anti-id mabs was confirmed by testing their binding to 18C2 hybridoma cells in the presence of homologous and heterologous "cold" inhibitor mabs. Experiments were also conducted to determine the functional properties of these mabs. Anti-18C2(id) mab 12H1.C5 inhibited the cytotoxic activity of rat splenic NK (nylon wool nonadherent cells, NWNA) and rat ALAK cells. Flow cytometric (FCM) analysis of the binding of the anti-18C2(id) mabs demonstrated that mab 12H1.C5 bound 75.43% rat NWNA spleen cells, 43.74% rat ALAK cells, and 74.33% rat CRC- cells. Anti-id mab 6D9.B11 bound 45.20% NWNA cells, 70.45% rat ALAK cells, and 55.86% CRC- cells. Two-color FCM analysis demonstrated that the anti-id mabs not only bound to the same molecule on NK cells, but also these mabs bound to the same molecule as 5C6, an anti-NK cell mab. Biochemical analysis of the antigen recognized by mab 12H1.C5 was determined by Western blotting. The determinant on NWNA cells recognized by mab 12H1.C5 had an M(r) of 40 kDa and appeared to be identical to that recognized by mab 5C6. The same experiment using a transformed rat RNK-16 (CRC-) cell extract and Western blot analysis, demonstrated an M(r) of 42 and 48 kDa in the presence of mabs 5C6 and 12H1.C5. Monoclonal antibody 5C6 was previously shown to recognize a vimentin-like function-associated molecule on NK cell membranes. The anti-id mabs were also shown to have cross-reactivity with the intermediate filament vimentin as determined by Western blot analysis.

In-situ hybridization using biotinylated probes to human papillomavirus (HPV) DNA was performed on formalin fixed paraffin embedded tissue in 30 patients with histologically confirmed adenocarcinoma-in-situ (AIS). Thirteen of the 30 cases contained areas of endocervical glandular dysplasia (EGD) admixed with AIS. Twenty one patients showed positive staining of the AIS nuclei for HPV DNA. Ten cases (33%) were positive for HPV 16 DNA and 11 cases (37%) were positive for HPV 18 DNA. No case showed synchronous expression of HPV 16 and 18 DNA. All cases of AIS were negative for HPV 6b and 11 DNA. Four cases of EGD were positive for HPV 18 DNA and 2 cases were positive for HPV 16 DNA. Four of 6 cases of intestinal dysplasia/AIS were positive for HPV 18 DNA. Associated squamous abnormalities (HPV +/- CIN +/- SCC) were noted in 15 cases. Of these, 7 showed positive staining for HPV DNA in the squamous lesion. Moreover, 5 of these were positive in both the AIS and squamous lesion. In-situ hybridization using biotinylated DNA probes is a sensitive and safe technique readily adaptable to routine histopathology.

Human immunodeficiency virus (HIV) infects cells after binding of the viral envelope glycoprotein gp120 to the cell surface recognition marker CD4. gp120 is noncovalently associated with the HIV transmembrane envelope glycoprotein gp41, and this complex is believed responsible for the initial stages of HIV infection and cytopathic events in infected cells. Soluble constructs of CD4 that contain the gp120 binding site inhibit HIV infection in vitro. This is believed to occur by competitive inhibition of viral binding to cellular CD4. Here we suggest an alternative mechanism of viral inhibition by soluble CD4 proteins. We demonstrate biochemically and morphologically that following binding, the soluble CD4 proteins sT4, V1V2,DT, and V1[106] (amino acids 1-369, 1-183, and -2 to 106 of mature CD4) induced the release of gp120 from HIV-1 and HIV-1-infected cells. gp120 release was concentration-, time-, and temperature-dependent. The reaction was biphasic at 37 degrees C and did not take place at 4 degrees C, indicating that binding of soluble CD4 was not sufficient to release gp120. The appearance of free gp120 in the medium after incubation with sT4 correlated with a decrease in envelope glycoprotein spikes on virions and exposure of a previously cryptic epitope near the amino terminus of gp41 on virions and infected cells. The concentration of soluble CD4 proteins needed to induce the release of gp120 from virally infected cells also correlated with those required to inhibit HIV-mediated syncytium formation. These results suggest that soluble CD4 constructs may inactivate HIV by inducing the release of gp120. We propose that HIV envelope-mediated fusion is initiated following rearrangement and/or dissociation of gp120 from the gp120-gp41 complex upon binding to cellular CD4, thus exposing the fusion domain of gp41.

A detailed study was performed in 14 patients with epithelial ovarian tumours using the satellite probes 33.15, 228S and 216S to investigate the nature of somatic changes and frequency with which clonal changes could be demonstrated during metastasis and progression. Somatic changes were evident in approximately 70% of ovarian tumours, the most common being a deletion or reduction in intensity of a band suggesting loss of heterozygosity. Additional changes that were observed included increased intensification of single bands and the appearance of novel DNA fragments. Somatic alterations were seen following digestion of DNA with methylation resistant restriction endonucleases indicating that methylation differences alone could not account for all of the somatic changes. Using DNA fingerprint analysis ovarian tumours were shown to be heterogeneous with different DNA patterns observed in different sites in five of eight patients. Generally, within an individual patient the primary and metastases appeared to share a DNA fingerprint pattern with minor variations occurring in different sites suggesting that different populations have derived from a common stem line. This study clearly demonstrates that DNA fingerprint analysis is a sensitive method to detect somatic changes in tumour DNA and for investigating the development of clonal heterogeneity in ovarian tumours.

A sensitive immunocytochemical technique was developed to detect a 170,000 dalton cell membrane glycoprotein (P-gp) in cell lines resistant to vincristine and vinblastine with varying degrees of resistance. P-gp was shown very clearly using the C219 monoclonal antibody and immunocytochemical detection with either antialkaline phosphate or peroxidase-antiperoxidase with silver gold intensification. There was good correlation between the results obtained with immunocytochemical detection of P-gp in single cells and Western blot analysis. The technique is easily performed and can detect P-gp in relatively small numbers of cells that Western blot analysis could miss and is suitable for clinical application.

Human hybrid tumor cells have been produced by fusing cells from freshly harvested tumor specimens with cells from a cultured human tumor line, D98OR. Fusions were performed with cells from 67 tumors and continuously growing hybrid lines were obtained from 16 (24%). A successful fusion usually produced 1 or 2 hybrid lines, but four easily fusable tumors produced from 6 to 26 lines. The parent cells and hybrids were analyzed by flow cytometry. Hybrids appeared to retain a high percentage of parental deoxyribonucleic acid. Ten patients participated in a clinical study in which they received intradermal immunization with semiautologous hybrids and Corynebacterium parvum as adjuvant. The only side effect was slight local tenderness at the injection sites. No tumor regressions occurred. Skin testing with parental and hybrid cells was performed prior to and following immunization with hybrids. Delayed cutaneous hypersensitivity was often achieved for hybrids but not for autologous tumor cells.