Search PubMed⌕ Search

Biomedical subjects

L W Stanton

Publications and source records attributed to L W Stanton.

36 records · Page 2Linked to original sources
PubMed1984

Neutralizing monoclonal antibodies to hepatitis A virus: partial localization of a neutralizing antigenic site.

BALB/c mice were immunized with purified preparations of hepatitis A virus (HAV) isolated after 21 days of growth in LLC-MK2 cells. The HAV antigen was isolated from CsCl gradients and consisted primarily of the following three proteins as analyzed after sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Coomassie blue staining: VP-1 at 33,000 daltons, VP-2 at 29,000 to 30,000 daltons, and VP-3 at 27,000 daltons. The spleen cells isolated from two BALB/c mice, immunized with two inoculations of HAV, were fused with SP 2/0 myeloma cells and grown in hypoxanthine-aminopterin-thymidine medium. Of 270 hybridomas initially screened, 72 were positive for binding HAV by a noncompetitive radioimmunoassay. All 72 were tested for the ability to neutralize the infectivity of HAV in an in vitro cell culture assay that was adapted for microtiter plates and that used detergent-treated virus for improved neutralization sensitivity and newborn cynomolgus monkey kidney cells for rapid growth. Eighteen hybridomas were positive for neutralization; 16 remained stable. Of the 16, 9 were able to compete with labeled polyclonal serum for binding to HAV. The nine competing hybridomas could be separated into two groups which appear to be directed towards two different sites on HAV and could complement each other in the competitive radioimmunoassay against polyclonal sera. Of the original 16 neutralizing hybridomas, 4 were subcloned through two cycles of limit dilutions. All four monoclonal antibodies retained their original neutralizing and competitive properties; three were immunoglobulin G2a, and one was immunoglobulin G1. All four monoclonal antibodies readily precipitate whole 125I-labeled HAV but are not able to recognize the disrupted proteins of the virus (as tested by immune precipitations of heat- and detergent-disrupted virions or Western blot analyses). However, the heterobifunctional cross-linking reagent toluene-2,4-diisocyanate was used to cross-link purified Fab fragments of two different monoclonal antibodies (2D2 and 6A5) to HAV before disruption. This reagent demonstrated a specific reaction of the monoclonal antibodies to the VP-1 of HAV, suggesting this major surface protein contains at least one of the major neutralization sites for HAV.

Animals↗
PubMed1983

Transcriptionally active c-myc oncogene is contained within NIARD, a DNA sequence associated with chromosome translocations in B-cell neoplasia.

NIARD (non-immunoglobulin-associated rearranging DNA) is located on mouse chromosome 15 at the break point of a commonly observed translocation event involving chromosomes 15 and 12 in murine plasmacytomas. The human cellular analogue of the v-myc oncogene of avian myelocytomatosis virus, strain MC-29, is known to reside on the distal end of human chromosome 8 and has been observed to translocate to chromosome 14 in Burkitt lymphomas. Using a cDNA clone specific for the transcript of the human c-myc gene (H c-myc), we show that the mouse c-myc (M c-myc) gene is contained within NIARD. NIARD-associated chromosome translocations occurred 1.3-2 kilobases (kb) 5' of the mouse c-myc gene where NIARD recombines with the switch region of the C(alpha) immunoglobulin gene in various murine plasmacytomas. The mouse c-myc encoding region within NIARD spanned <2.4 kb of DNA and expressed a low level of a 2.3-kb polyadenylylated RNA in BALB/c spleen. Increased (10- to 20-fold) levels of rearranged mouse c-myc transcripts (i.e., approximately 1.8-2.1 kb) were observed in plasmacytomas that have NIARD-associated chromosome translocations. Human c-myc and NIARD probes detected DNA rearrangements of human c-myc in four of seven Burkitt lymphomas. DNA sequences adjacent to the human c-myc gene recombined with the C(mu) immunoglobulin gene locus on chromosome 14 in several Burkitt lymphomas. The activation of the c-myc oncogene by chromosome translocation implicates its involvement in B-cell oncogenesis.

Animals↗
PubMed1982

Nucleotide sequence and properties of the murine gamma 3 immunoglobulin heavy chain gene switch region: implications for successive C gamma gene switching.

During B lymphocyte differentiation, immunoglobulin heavy chain constant region (CH) genes undergo a unique series of DNA recombination events culminating in the CH class switch. CH switch (S) regions are located 2 kb 5' of each CH gene except delta (i.e. mu, gamma 3, gamma 1, gamma 2b, gamma 2a, epsilon and alpha). We describe the structural features of the gamma 3 switch region. Hybridization experiments show that S gamma 3 has remarkable homology to both S mu and other S gamma regions while S mu possesses limited homology to the other S gamma sequences. However, S mu possesses extensive sequence homology with S epsilon and S alpha. The nucleotide sequence of S gamma 3 reveals higher densities of S mu repetitive sequences (GAGCT and GGGGT) and another S region common sequence (YAGGTTG) than observed for S gamma 1, S gamma 2b or S gamma 2a. In addition, the conservation of S mu like repetitive sequences in S gamma regions is correlated with the 5' leads to 3' gamma gene order (i.e. S gamma 3 greater than S gamma 1 greater than S gamma 2b greater than S gamma 2a). A model is presented which suggests that the unique features of S gamma 3 may allow for successive switches from C mu to any C gamma gene.

Animals↗
PubMed1982

On immunoglobulin heavy chain gene switching: two gamma 2b genes are rearranged via switch sequences in MPC-11 cells but only one is expressed.

During B lymphocytes differentiation, switches in the expression of heavy chain immunoglobulin constant region (CH) genes occur by a novel DNA recombination mechanism. We have investigated the requirements of the CH gene switch by characterizing two rearranged gamma 2b genes from a gamma 2b producing mouse myeloma (MPC-11). One of the two gamma 2b genes is present in 2-3 copies per cell (gamma 2b strong hybridizer) while the other is present in approximately 1 copy per cell (gamma 2b weak hybridizer). Genomic clones of the gamma 2b strongly hybridizing gene indicate that this is an abortive switch event between the S gamma 3 and S gamma 2b regions. However, clones of the gamma 2b weakly hybridizing gene suggest a functional rearrangement due to the presence of VH, JH and S mu sequences. The switch-recombination sites of these rearranged gamma 2b genes and those of other CH genes show a high degree of preference for the sequence AGGTTG 5' of either the S mu donor site or the appropriate CH S acceptor site. AGGTTG and its analogs are rare in the S mu region, are somewhat prevalent in s alpha and in the case of S mu are found 5' of a tandemly repeated DNA sequence (GAGCT, GGGGT) comprising most of S mu.

Animals↗
PubMed1981

Restriction endonuclease analysis of the DNA from varicella-zoster virus: stability of the DNA after passage in vitro.

DNA was extracted from nucleocapsids isolated from WI-38 cells infected with two different strains of varicella-zoster virus (VZV). The DNAs were treated with each of six restriction endonucleases (EcoRI, HindIII, Bg/I, Bg/II, Sal I and Bam-HI) and small, but reproducible differences in restriction endonuclease patterns were observed. These strains were passaged in WI-38 cells and in primary guinea-pig embryo (GPE) cells, followed by restriction endonuclease analysis of the DNAs. No changes were observed in the restriction prpofile of the DNA of one of the strains (VZV-KMcC) after 46 passages in WI-38 cells but small differences were observed after 72 passages. No changes were observed after 30 passages of another strain (VZV-AW) in WI-38 cells. Twenty passages of VZB-KMcC in GPE cells did result in minor alterations of its DNA. It was concluded that VZV DNA was sufficiently stable after multiple passages in WI-38 cells to make restriction endonuclease analysis a valuable epidemiological tool for strain differentiation.

Animals↗
PubMed1981

Immune responses in mice against herpes simplex virus: mechanisms of protection against facial and ganglionic infections.

We performed experiments with mice to determine the nature of the immune response(s) that prevents primary infections of the skin and the trigeminal ganglia with herpes simplex virus. Immunization with infectious herpes simplex virus, inactivated virus, or material enriched for viral glycoproteins protected hairless mice against primary facial and ganglionic infections. Live and inactivated viruses induced neutralizing antibodies, whereas glycoprotein material did not. Instead, glycoprotein material induced antibodies that were largely directed against two glycopolypeptides with molecular weights of 120,000 to 130,000. Hairless mice immunized with glycoprotein material responded faster than control mice in the synthesis of neutralizing antibodies after challenge with infectious virus. Congenital athymic BALB/c (nu/nu) mice were protected against primary facial infections after immunization with glycoprotein material, but glycoprotein-specific antibodies were not induced.

Animals↗
PubMed1981

Immune response to herpes simplex virus infections: virus-specific antibodies in sera from patients with recurrent facial infections.

Radioimmunoprecipitation assays were used to identify antibodies against a number of herpes simplex virus type 1-specific antigens in serum samples from individuals with recurrent facial herpes virus infections and from seropositive individuals without recurrent infections. Individuals with recurrent infections contributed three sequential serum samples each: immediately after the appearance of lesions, 3 weeks later, and 3 months later. Antibodies against at least 18 viral polypeptides were present in all positive sera: these included antibodies against the major nucleocapsid polypeptide (approximate molecular weight, 150,000) and against two glycopolypeptides with molecular weights of 115,000 to 130,000. No significant differences were observed between the serum samples in regard to their virus-specific antibody composition. The high-molecular-weight glycopolypeptides were partially purified and used in quantitative titration experiments. All sera tested were equally reactive with this material. It was concluded that under the experimental conditions an individual's susceptibility to recurrent herpetic infections could not be correlated with quantitative or qualitative changes in the levels of virus-specific antibodies.

Antibodies, Viral↗
PubMed1980

Percutaneous transluminal dilatation of the renal arteries: techniques and results.

The techniques and results of percutaneous transluminal angioplasty (TPA) of the renal arteries were evaluated in 20 hypertensive patients with 25 renal artery stenoses. Follow-up angiograms have been obtained in nine patients. Many patients had evidence of both essential and angiotensinogenic hypertension. Eleven patients had evidence of severe diffuse atherosclerotic disease, and nine patients had evidence of renal insufficiency. The mean systolic pressures before and after PTA were 203.80 and 150.30, respectively. The mean diastolic pressures before and after PTA were 117.45 and 85.95, respectively. Ten patients were cured. Six patients with long histories of hypertension and a recent increase in blood pressure were classified as having blood pressure easier to control with antihypertensive medication following PTA. Three patients failed to respond to PTA, and one patient was a technical failure. The advantages of this technique include avoidance of general anesthesia and a major surgical procedure, decreased cost, and a shortened hospital stay. The technique can be easily repeated if necessary, and future surgical intervention is not precluded if the method is unsuccessful.

Adult↗
PubMedDate not supplied

Translocation, breakage and truncated transcripts of c-myc oncogene in murine plasmacytomas.

Comparative nucleotide sequence analysis of a rearranged c-myc gene in a murine plasmacytoma and c-myc cDNA from normal spleen reveals that chromosomal translocation in the plasmacytoma breaks the c-myc gene within the first exon or intron. In the plasmacytoma truncated c-myc RNAs initiate from newly exposed promoter sites. Nevertheless, the myc polypeptide produced in the plasmacytoma is probably the same as that from the intact c-myc gene because the exon lost by breakage and translocation is non-coding. The second and third exons of the mouse c-myc gene are substantially conserved in the v-myc gene of the avian retrovirus, MC29.

Animals↗
PubMedDate not supplied

Nucleotide sequence of cloned cDNA of human c-myc oncogene.

Like other transforming genes of retroviruses, the v-myc gene of the avian virus, MC29, has a homologue in the genome of normal eukaryotic cells. The human cellular homologue, c-myc, located on human chromosome 8, region q24 leads to qter (refs 1, 2), is translocated into the immunoglobulin heavy-chain locus on human chromosome 14 (ref. 3) in Burkitt's lymphoma, suggesting that c-myc has a primary role in transformation of some human haematopoietic cells. In addition, c-myc is amplified in the human promyelocytic leukaemia cell line, HL60 (refs 6, 7) which also contains high levels of c-myc mRNA. Recently, Colby et al. reported the nucleotide sequence of the human c-myc DNA isolated from a genomic recombinant DNA library derived from human fetal liver. This 4,053-base pair (bp) sequence includes two exons and one intron of the myc gene, and the authors have suggested the existence of a human c-myc mRNA of 2,291 nucleotides that has a coding capacity for a protein of molecular weight (Mr) 48,812. We have approached the problem of accurately defining the characteristics of the human c-myc mRNA and c-myc protein by determining the sequence of the c-myc cDNA isolated from a cDNA library prepared from mRNA of a clone of the K562 human leukaemic cell line. K562 cells are known to contain c-myc mRNA which is similar in size to the c-myc mRNA of other human cell types. We report here the sequence of 2,121 nucleotides of a human c-myc mRNA and demonstrate that its 5' noncoding sequence does not correspond to the sequence of the reported genomic human sequence. However, our data confirm that the intact human c-myc mRNA can encode a 48,812-Mr protein with a sequence identical to that reported by Colby et al.

Amino Acid Sequence↗
PubMedDate not supplied

Nucleotide sequence comparison of normal and translocated murine c-myc genes.

Plasmacytoid tumours in mice and Burkitt's lymphomas in humans display characteristic chromosomal translocations involving the c-myc proto-oncogene. Models to explain quantitative changes in c-myc expression have been proposed based on the loss of normal promoters as a result of translocation. However, alternative explanations, such as somatic mutation are needed to explain altered c-myc expression in the absence of gene breakage. We present here the nucleotide sequence of the normal murine c-myc gene. Comparison of this sequence with that of a translocated c-myc gene from a murine plasmacytoma reveals complete identity of coding sequence. One nucleotide difference was found in the non-coding first exon. This shows that qualitative changes of the c-myc gene product are not required for oncogenesis in murine plasmacytomas. In contrast, mutations are found in coding and non-coding regions of translocated c-myc genes from Burkitt's lymphomas, suggesting that the mechanisms by which c-myc is activated in plasmacytomas and Burkitt's lymphomas are different.

Amino Acid Sequence↗