Biomedical subjects

D Wojtacha

Publications and source records attributed to D Wojtacha.

6 recordsLinked to original sources

PubMed2006

Fluorescence-activated single cell sorting of human embryonic stem cells.

Human embryonic stem cells (hESC) are the subject of intense investigation for use in regenerative medicine, in toxicity testing, and as models for the study of human development. Automated cell sorting will enhance the isolation of homogenous pools of differentiated hESCs both for basic studies and for therapeutic applications. Sorting could also be used to deplete undifferentiated, potentially tumourigenic cells. However, hESCs are sensitive to single cell disaggregation and recover poorly when plated at clonal density. Here we report a method for successful semi-automated single cell sorting of hESCs. This method utilizes an ES-specific promoter-transgene construct and automated FACS-based single cell sorting and plating. Clonal recovery in physiologic oxygen (2%) was increased fourfold over room oxygen (21%; p < 0.01). This automated protocol will help to realize proposed hESC strategies that are hampered by low throughput and poor yields.

Alkaline Phosphatase↗

PubMed1994

Expression of tissue kallikrein in human kidney.

1. We studied the distribution of human tissue kallikrein mRNA in normal and diseased kidney, using in situ hybridization, together with immunohistochemical localization of renal kallikrein protein. Materials studied were (a) normal tissue from kidneys removed because of localized renal carcinoma, (b) kidneys removed because of post-traumatic haemorrhage and (c) renal biopsy specimens from patients with membranous glomerulonephritis and nephrotic syndrome. 2. A 1.35 kb EcoRI fragment of human tissue kallikrein cDNA was labelled with [32P]dCTP using the random-primer technique, and used for in situ hybridization. A specific rabbit antibody to active human urinary kallikrein was employed for immunocytochemistry, using a peroxidase-antiperoxidase method. 3. By in situ hybridization, no tissue kallikrein gene expression was seen in the carcinoma nephrectomy specimens. Positive expression was seen in the trauma nephrectomy tissue, and in four of five nephrotic syndrome biopsies. In all kidneys, expression was confined to the renal cortex. The dominant site of gene expression was the distal tubule. Apart from one area of positive signal related to an epithelial cell of Bowman's capsule, expression was not observed in glomeruli. Expression was also seen in the walls of large- and medium-sized blood vessels. 4. By immunohistochemistry, the dominant site of immunoreactivity was the distal tubule. Dense staining was also seen in granular peripolar cells and in isolated parietal epithelial cells close to the vascular pole. Isolated immunoreactive cells were seen in the media of large- and medium-sized arteries. 5. The tissue kallikrein gene in the kidney may not be constitutively expressed, but is expressed in response to physiological or pathological stimuli.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1992

Detection of cytomegalovirus antigens in phagocytosed serum complexes from a patient with rheumatoid arthritis, vasculitis, peripheral neuropathy, cutaneous ulceration, and digital gangrene.

A patient with rheumatoid arthritis, vasculitis, peripheral neuropathy, cutaneous ulceration, and digital gangrene was studied. Circulating immune complexes were detected by C1q binding although serum complement levels were within the normal range. Immunofluorescent staining of buffy coat cells with specific antisera showed the presence of IgG and IgM in phagocytosed inclusions but complement C3 was not detected. A monoclonal antibody specific for cytomegalovirus detected antigens in phagocytosed inclusions on one occasion. These results may suggest that cytomegalovirus antigens are a hitherto unidentified component of serum complexes in patients with rheumatoid arthritis and may contribute to the pathogenesis of the vasculitic complications of rheumatoid arthritis by participating in immune complex formation.

Antigen-Antibody Complex↗

PubMed1990

In situ hybridisation.

In situ hybridisation of mRNA in tissues or cell preparations is a powerful technique for studying gene expression. When combined with cell phenotyping with monoclonal antibodies it gives insights into the cellular basis of disease in vivo. The technique has also been used widely to identify foreign nucleic acids--for example, bacterial or viral, in host cells. The major disadvantages of this approach in the past have been that it was technically demanding, time consuming, and provided qualitative rather than quantitative results. Now, with the use of non-radioactive probes and improved imaging systems, the full potential of this form of molecular analysis is increasingly accessible and should generate rapid advances in many fields.

Arthritis, Rheumatoid↗

PubMed1988

Do polyclonal rheumatoid factors carry an 'internal image' of cytomegalovirus, Epstein-Barr virus and nuclear antigens?

Rabbit antisera have been prepared against isolated rheumatoid factors (RF's). It was considered that RFs are anti-idiotype antibodies and that the anti-RF antisera had anti-anti-idiotype specificity. Furthermore, it was considered that the RF's carry an "internal image" of the putative "antigen X" and that the rabbit antisera would have specificity for this "antigen-X". Reactions of the rabbit antisera with the early and late antigens of CMV, EBV antigens and nuclear antigens suggest that there may be an "internal image" of these antigens in rheumatoid factor molecules and that they all may be related to the immunogenesis of RF.

Animals↗

PubMed1978

Immunological studies of the placenta in systemic lupus erythematosus.

An immunological study was made of the placentae from 5 mothers with lupus erythematosus. 3 of the 5 mothers had anti-DNA antibodies in their sera at the time of delivery and in one of these anti-DNA antibodies were detected in the cord blood. This patient had active renal disease and serological evidence suggestive of circulating immune complexes in her blood at the time of delivery. Immunofluorescence studies showed granular deposition of immunoglobulin and C3 on the trophoblast basement membrane similar to that previously described on the glomerular basement membrane in systemic lupus erythematosus. Anti-DNA antibodies were eluted from the placenta in this case. We suggest that immune complex deposition on the trophoblast basement membrane in patients with active systemic lupus erythematosus may play a part in the increased fetal mortality in this disease.

Adult↗