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Biomedical subjects

G Mitra

Publications and source records attributed to G Mitra.

At least 37 records · Page 2Linked to original sources
PubMed1989

Virus inactivation/elimination in therapeutic protein concentrates.

Concern for virus transmission from intravenous biological products, derived from natural and biotechnological sources, has prompted us to investigate various methods for virus inactivation while preserving the biological activity of the therapeutic entity. This concern is currently acute regarding the presence of HIV in purified human plasma protein concentrates. Four different methods of virus removal (1) heating in solution; (2) use of lipid solvents; (3) low pH treatment, and (4) partitioning during purification, are reported for various plasma proteins that have been spiked with model viruses. General virus inactivation strategies for therapeutic protein concentrates are discussed.

Animals↗
PubMed1988

Inactivation of viruses in therapeutic products derived from human plasma.

Alpha-1-proteinase inhibitor concentrates have been prepared from pooled human plasma for therapeutic applications. Pasteurization (60 degrees C, 10 hours) conditions have been defined to reduce risks of transmission of viral agents without significant loss of biologic activity of the purified product. Human clinical data collected to date support the model virus inactivation studies regarding viral safety.

Blood Proteins↗
PubMed1987

Characterisation of various intravenous immunoglobulin preparations.

Five commercially available immunoglobulin preparations are characterised with regard to molecular weight distribution, immunoglobulin subclass distribution and antibody content. Preparations processed utilising enzymic or chemical modification demonstrate considerable variance of molecular weight species and subclass distribution when compared to native preparations. Similarly, cytomegalovirus and anti-streptolysin O antibody content is dramatically reduced in the modified preparations. Possible reasons for the differences are discussed. In-process inactivation of Human Immunodeficiency Virus is reviewed with regard to one of the native preparations.

Antibodies, Viral↗
PubMed1987

Identification and biochemical characterization of p70TRK, product of the human TRK oncogene.

TRK is a human transforming gene generated in a colon carcinoma by a somatic rearrangement that fused a nonmuscle tropomyosin gene to sequences that shared extensive homology with members of the tyrosine-protein kinase supergene family. These sequences are likely to be derived from a transmembrane receptor gene whose putative ligand binding domain has been replaced by tropomyosin. In the present studies, we have expressed the entire coding sequences of the TRK oncogene as well as its protein kinase-related carboxyl-terminal domain in Escherichia coli. Antisera raised against these bacteria-synthesized TRK polypeptides has allowed us to identify the gene product of the TRK oncogene as a 70-kDa protein. Immunoprecipitates containing p70TRK have an associated protein kinase activity specific for tyrosine residues. Moreover, p70TRK is phosphorylated in vivo in serine (75%), threonine (20%), and tyrosine (5%) residues. Finally, immunofluorescence and cellular fractionation studies indicate that p70TRK is preferentially located in the cytoplasmic fraction.

Amino Acids↗
PubMed1986

Monoclonal antibody binding to factor VIII:c.

The interactions of human blood clotting Factor VIII:c with 2 monoclonal antibodies, C7F7 and BD10, were examined with an enzyme-linked immunosorbent assay (ELISA) method. A Scatchard-Sips plot approach to data analysis was used to calculate an average affinity (Ko), valence (n), and heterogeneity index (a) for each interaction. Using a Factor VIII:c coating ELISA method, the following constants were determined: BD10: Ko = 5.48 X 10(8) M-1; n = 0.31; a = 0.97; C7F7: Ko = 2.3 X 10(11) M-1; n = 0.018; a = 0.77.

Antibodies, Monoclonal↗
PubMed1986

Preparation of high purity factor VIII concentrates.

An improved process for producing high purity Factor VIII concentrate has been developed. The improvement was based on the combination of three precipitating agents (polyethylene glycol, glycine and sodium chloride) which heretofore have been used alone in a single step or in separate and distinct steps in a multi-step process. The product was pasteurized in solution (60 degrees C, 10 hrs) to reduce possible risks of virus transmission. The purified product has significantly higher specific activity and lower fibrinogen and immunoglobulin levels when compared to other commercial concentrates.

Chemical Precipitation↗
PubMed1986

Hepatitis B virus associated DNA polymerase inactivation in factor IX concentrates.

Inactivation of Hepatitis B virus associated DNA polymerase was studied in factor IX concentrate (coagulation factors II, VII, IX and X) by heat pasteurization (60 degrees C, 10 hr) and by alkylating agents iodoacetic acid and iodoacetamide. DNA polymerase appeared to reach a residual level which occurred in human serum albumin at 60 degrees C, 10 hr under comparable spike level of hepatitis B virus. Of the four coagulation factors, factor IX activity was most susceptible to inactivation procedures with 40-50% recovery across heat pasteurization and approximately 70% recovery across iodoacetic acid treatment. Factor IX specific activities of the treated concentrates were greater than or equal to 70% of the untreated controls with no appreciable change of corresponding NAPTT values. Factor IX concentrates subjected to such inactivation procedures should reduce the potential for hepatitis B virus transmission.

Blood Coagulation Factors↗
PubMed1986

The effect of lead on insecticide metabolism.

The possibility that lead could affect the metabolism of the insecticide 1,1-dichloro-2, 2-bis(p-chlorophenyl)ethane, DDD, was examined by studies of the effects of chronic oral Pb treatment on DDD conversion to 2,2-bis(p-chlorophenyl)acetic acid, DDA. Rats were given either distilled deionized water or 0.05, 0.58, 17 or 352 ppm Pb solutions as drinking water. Systolic blood pressure and body weight were measured weekly. Rats drinking 352 ppm lead chloride manifested a statistically significant increase in blood pressure. Rats drinking 352 ppm and 17 ppm lead chloride showed a significant decrease in the rate of weight gain compared to controls. All groups showed an increase in the excretion of total DDA compared to controls.

Animals↗
PubMed1985

Saccharomyces cerevisiae coordinates accumulation of yeast ribosomal proteins by modulating mRNA splicing, translational initiation, and protein turnover.

The rate of accumulation of each ribosomal protein is carefully regulated by the yeast cell to provide the equimolar ratio necessary for the assembly of the ribosome. The mechanisms responsible for this regulation have been examined by introducing into the yeast cell extra copies of seven individual ribosomal protein genes carried on autonomously replicating plasmids. In each case studied the plasmid-borne gene was transcribed to the same degree as the genomic gene. Nevertheless, the cell maintained a balanced accumulation of ribosomal proteins, using a variety of methods other than transcription. (i) Several ribosomal proteins were synthesized in substantial excess. However, the excess ribosomal protein was rapidly degraded. (ii) The excess mRNA for two of the ribosomal protein genes was translated inefficiently. We provide evidence that this was due to inefficient initiation of translation. (iii) The transcripts derived from two of the ribosomal protein genes were spliced inefficiently, leading to an accumulation of precursor RNA. We present a model which proposes the autogenous regulation of mRNA splicing as a eucaryotic parallel of the autogenous regulation of mRNA translation in procaryotes. Finally, the accumulation of each ribosomal protein was regulated independently. In no instance did the presence of excess copies of the gene for one ribosomal protein affect the synthesis of another ribosomal protein.

Cloning, Molecular↗
PubMed1984

Recovery and inactivation of infectious retroviruses from factor VIII concentration.

The ability of infectious retroviruses to withstand the procedures used for factor VIII concentration was investigated. Mouse retroviruses added to human plasma survived these procedures and remained infectious in lyophilised samples of factor VIII. Lyophilised material had to be heated at 68 degrees C for several hours before substantial quantities of infectious virus became inactivated. These findings support the possible role of retroviruses in AIDS, and indicate that factor VIII concentrates must be heated to inactivate these infectious viruses.

Animals↗
PubMed1984

A yeast ribosomal protein gene whose intron is in the 5' leader.

The complete gene encoding yeast ribosomal protein 29, a component of the 60 S ribosomal subunit of Saccharomyces cerevisiae has been isolated and its nucleotide sequence determined. The coordinates of transcription initiation and termination of the rp29 gene have been mapped. Transcription appears to start at several sites, spanning nearly 30 nucleotides. The gene is transcribed into a precursor RNA molecule of 1110 to 1085 nucleotides from which an intron of 458 nucleotides is spliced out. The splice junction has been determined by comparing the nucleotide sequences of the rp29 gene with those of a homologous cDNA clone. Interestingly, the intron of the rp29 gene resides in the 5' untranslated region of the rp29 mRNA, the first instance where this has been observed in yeast. The single open reading frame in the rp29 gene encodes a very basic protein of 155 amino acid residues.

Amino Acid Sequence↗
PubMed1984

Genetic structure and stability of a copy number mutant of IncFI group plasmid ColV-K94 in Escherichia coli K-12.

Mutants pWS10, pWS11 and pWS12 were derived from an IncFI group plasmid ColV-K94 by the insertion of a transposon Tn903 (Kmr). These plasmids were all approximately 130 kb in length. The plasmid pWS12 resembled the wild type ColV-K94 in transmissibility, incompatibility and stable maintenance. Cells harboring pWS11 were poor conjugal donors but resistant to the same level of kanamycin as pWS12 containing hosts. In contrast, pWS10 conferred a higher resistance to kanamycin and exhibited reduced incompatibility properties in comparison with pWS12. The higher drug resistance associated with pWS10 appeared to be a consequence of an increase in its copy number and the generation of miniplasmids of varying sizes. Electron microscope analysis of the copy mutant pWS10 revealed that Tn903 was located at a site adjacent to a region 32.6F to 35.3F. The latter region appears to be the primary replicon of ColV-K94 and is homologous with the secondary replicon of F. The insertional mutagenesis with Tn903 brought about an extensive DNA rearrangement including the duplication and translocation of the stems of two inverted repeat structures. The DNA alterations of pWS10 were distinguishable through comparison of its EcoRI digestion patterns with those of pWS11 and pWS12.

DNA Transposable Elements↗
PubMed1984

Stabilization of alpha-1-proteinase inhibitor by citrate.

alpha 1-Proteinase inhibitor has potential therapeutic value in patients with chronic deficiency (which can lead to emphysema) and in acute adult respiratory distress syndrome. Pasteurization (10 h, 60 degrees C) can reduce the risk of hepatitis B; and addition of stabilizers to the inhibitor helps prevent the loss of biological activity which occurs during pasteurization. Denaturation during heating at several temperatures was studied, using high pressure liquid chromatography, differential scanning calorimetry, and elastase inhibitory capacity to measure the extent of denaturation. The presence of 0.5 M citrate increases the Arrhenius-type activation energy 3-fold, and the enthalpy of denaturation is increased by 3.5 kJ/mol (15 kcal/mol). The onset temperature of the endotherm is increased by more than 10 degrees C. The inhibitor is further stabilized by sucrose (O.5 g/ml). In the presence of both citrate and sucrose, samples of alpha 1-proteinase inhibitor retained almost full biological activity following pasteurization for 10 h at 60 degrees C; the heating conditions used to destroy hepatitis B infectivity of other plasma derivatives.

Blood Proteins↗
PubMed1984

Second replicon in ColV2-K94 mediates the stable coexistence of two incompatible plasmids.

The colicin-producing plasmid pWS12, a Tn903 derivative of ColV2-K94, was found to be incompatible with the IncFI plasmids KLF1 and R386. It was compatible with the IncFII plasmids R538 and R100. Three overlapping mini-ColV derivatives, pWS15, pWS16 and pWS17, were obtained by restriction digestion of pWS12. Unlike pWS12, pWS16 exhibited incompatibility with both IncFI and IncFII plasmids, whereas the pWS15 and pWS17 plasmids expressed IncFII incompatibility but not the IncFI incompatibility of their parental ColV plasmid. We show that, although pWS12 has an IncFII replicon, Rep1, it does not normally express IncFII incompatibility because a second replicon, Rep2 (homologous to the secondary replicon of F), functions during the stable coexistence of the plasmid with IncFII plasmids. When Rep2 is deleted (as in the mini-ColV plasmids) or made nonfunctional (as in a PolA mutant strain), ColV then behaves as an IncFII plasmid.

Bacteriocin Plasmids↗
PubMed1981

Studies on ultrafiltration of antihemophilic factor.

Ultrafiltration as an alternative approach to reprecipitation was studied in the production of an intermediate-purity antihemophilic factor concentrate suitable for clinical use. Almost complete recoveries of factor-VIII-related activities have been demonstrated across the processing step resulting in increased net yield of the product. Several important processing parameters are discussed. The product thus prepared is stable and has improved VIII:C/mg fibrinogen ratio.

Electrophoresis, Cellulose Acetate↗
PubMed1978

Continuous fractionation of human plasma.

Continuous processing has been applied to human plasma fractionation by the cold ethanol process. On-line pH control of +/- 0.05 pH units, flow control of +/- 1%, and temperature control of +/- 0.5 degree C have been achieved. Optimization of precipitation pHs has been carried out for purifying plasma protein fractions and albumin. During precipitation, the irreversible nature of the pH overshoots has been demonstrated. Compared to the batch processing mode, the continuous scheme produces an increased yield between 6 to 11%.

Autoanalysis↗