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Biomedical subjects

R C Briggs

Publications and source records attributed to R C Briggs.

At least 55 records · Page 3Linked to original sources
PubMed1984

Two human colon tumor cell lines with similar nuclease sensitivities have different ethidium bromide binding characteristics.

Chromatin from two human colon adenocarcinoma cell lines (HT-29 and LoVo) showed similar digestion kinetics when sensitivities to DNase I and micrococcal nuclease were examined. Chromatin conformations were probed by examining the binding of ethidium bromide. A Scatchard plot revealed that both chromatins bound the same amount of ethidium bromide per mole of DNA, but the DNA from LoVo cells was more accessible to the intercalator. The results indicate that differences in chromatin conformation are not necessarily accompanied by different nuclease sensitivities.

Adenocarcinoma↗
PubMed1984

Nonhistone protein antigen profiles of five leukemic cell lines reflect the extent of myeloid differentiation.

The human leukemic cell lines, K562, KG-1, and HL-60, and the blast subclones, KG-1a and HL-60 blast, were utilized to relate differences in nonhistone protein antigens to stages of myeloid cell differentiation. Chromatin proteins were separated on SDS-polyacrylamide gels, transferred electrophoretically to nitrocellulose sheets, and visualized by the peroxidase-antiperoxidase method of Sternberger. Screening with antisera raised against total and dehistonized chromatin and a nuclear extract from these cells revealed quantitative as well as qualitative differences between the cell lines. A decrease in antigen content seemed to parallel progressive stages of myeloid cell development. The results indicate that a number of chromosomal protein antigens are lost or modified during differentiation. An antigen(s) of approximately 55,000 molecular weight was found in HL-60 chromatin, but was not present in its less differentiated subclone or in the other lines representative of earlier stage cells. Upon the induction of HL-60 cells to mature to end stages with 4 microM retinoic acid, a significant increase in the mol wt 55,000 activity was seen. This antigen was detected only with antisera against HL-60 total chromatin and granulocyte nuclei, and it was found only in normal mature granulocytes and in the later stage cells of the HL-60 culture. Thus, the antigen appears to be associated with a differentiated myeloid function.

Antibodies, Neoplasm↗
PubMed1983

Crosslinking of chromosomal proteins to DNA in HeLa cells by UV gamma radiation and some antitumor drugs.

Immunochemical techniques were used to investigate the protein-DNA crosslinking by ultraviolet (UV) and gamma radiation as well as 1,3-bis-(2-chloroethyl)-1-nitrosourea (BCNU) or cis- and trans-diamminedichloroplatinum II (cis-DDP and trans-DDP). Antisera to 0.35 M NaCl extract and 0.35 M NaCl residue of HeLa nuclei were employed. Both gamma and UV irradiation, exposure to cis- or trans-DDP and, to a lesser extent, BCNU, resulted in crosslinking of various antigens to the DNA. Although several antigens were crosslinked by all the employed agents, other exhibited agent-specific crosslinking patterns.

Carmustine↗
PubMed1983

Tissue and cell-specific antigens in chromatin from cultured rat Sertoli cells.

Cell-specific chromatin antigens have been detected in rat Sertoli cells. Antisera were raised in rabbits to dehistonized chromatin prepared from 5- to 6-day cultures of rat Sertoli cells and immunoreactivity was assessed with microcomplement fixation tests or immunoidentification of antigens separated electrophoretically and transferred to nitrocellulose sheets. Tissue specificity was confirmed further by immunoabsorption. These antisera recognized only components of Sertoli cell chromatin; chromatins prepared from rat liver, kidney, thymus, Novikoff hepatoma, a testes germinal cell fraction, or purified rat DNA showed little or no immunoreactivity. Nitrocellulose transfers of total chromosomal proteins revealed the presence of two high-molecular-weight antigens (greater than 200,000) and a broad range of weaker immunologic species (M tau about 50,000-200,000) in Sertoli cell chromatin but not liver or thymus chromatin. Deproteinization of Sertoli cell chromatin with concentrated salt and urea at pH 6 or 8 produced an increase in complement-fixing activity of the fraction sedimenting with DNA and micrococcal nuclease digestion studies showed these fractions to depend in part on DNA for antigenicity. Individual antigens in the protein-DNA pellets of pH 8 salt and urea fractionated chromatin could not be identified with antigens on nitrocellulose sheets. Collectively, these observations suggest that at least two groups of cell-specific antigens exist in Sertoli cell chromatin; one group is detected after electrophoretic separation and transfer to nitrocellulose and the other group, reactive in complement fixation, cannot be detected through this method, apparently becoming antigenically inactive once the complex of protein and DNA is dissociated.

Animals↗
PubMed1983

Antigens in chromatin associated with proliferating and nonproliferating cells.

Xenoantisera were raised to total chromatin from the leukemia cell line K562, or materials released through limited deoxyribonuclease I digestion of nuclei or during the control incubation of nuclei without enzyme. The peroxidase-antiperoxidase method of antibody-antigen detection was employed to visualize individual antigens resolved on one-dimensional polyacrylamide gels following transfer to sheets of nitrocellulose (immunotransfers). Each antiserum contained multiple antigen specificities as evidenced by the diverse patterns of reactive bands displayed on the immunotransfers. The most striking difference in antigens recognized between the antisera was observed in the molecular weight region below 50,000, where two highly reactive bands were seen mainly with antiserum to nuclear materials released by deoxyribonuclease I digestion. The antigens detected with all of the antisera were present in chromatins prepared from proliferating cells, while the levels of antigens present in chromatin from non-proliferating peripheral blood lymphocytes were greatly reduced or not detected. Antigens in chromatin from proliferating cells that migrated with apparent molecular weights of 37,000 and 100,000 were not lost once the activities to antigens in lymphocyte chromatin were absorbed out. These two activities were absorbed from antisera with the same amount of chromatins from proliferating cells. Two antigens migrating at molecular weight 52,000 and 76,000 appeared more active in the chromatin from unstimulated lymphocytes than in chromatin from proliferating cells.

Cell Division↗
PubMed1983

Nuclear localization of lactoferrin in the human granulocyte: Artifact incurred during slide preparation.

Within the blood cells, lactoferrin is found only in the late stage neutrophilic granulocytes. Lactoferrin first appears in these cells during the myelocyte stage of development coincidentally with the specific or secondary granules. Most investigators report a cytoplasmic immunocytochemical localization reaction within the granulocyte. However, others have observed a prominent nuclear localization reaction. Treating the cells with certain fixatives was shown to prevent the relocation of lactoferrin from the cytoplasm to the nucleus when the localization was done on granulocytes prepared by smearing. The present study demonstrated that the relocation of lactoferrin is only a problem when cells were smeared or cytocentrifuged onto slides or fractionated for the purpose of isolating cellular organelles. Under these conditions the selection of fixative is an important consideration. Exposing isolated lactoferrin to a fixative effective in retaining lactoferrin in the cytoplasm of granulocytes smeared on slides did not alter a number of its physical properties. The results suggest that maintenance of the normal cytoarchitecture or effect of fixative on other cellular components prevents the relocation of lactoferrin within the cell during tissue processing and the direct action of fixation on lactoferrin is probably not responsible for this effect.

Acetone↗
PubMed1983

Rapid method for preparation of nuclei from a mucin-producing solid tumor.

Nuclei were prepared from the human colon mucin-producing tumor GW-39. The method described involves incubation in beta-mercaptoethanol under mild conditions and results in the preparation of nuclei free of the mucin web that initially enmeshes them. A sensitive colorimetric immunoassay was used to confirm that nuclei prepared in this fashion were generally free of contaminating mucin. This method is mild, rapid, and should be applicable to other mucin-producing tumors.

Cell Fractionation↗
PubMed1982

Immunological specificity of Novikoff hepatoma chromatin: isolation of three antigenic proteins.

1. Immunization of rabbits with dehistonized Novikoff hepatoma or normal rat liver chromatin elicited specific antibodies. 2. Chromosomal nonhistone proteins from Novikoff hepatoma were fractionated and three proteins (approx. mol. wt 39,000, 49,0000 and 56,000) were identified with the hepatoma antigenic complexes. 3. All three proteins showed significant reactivity of PAS staining suggesting that they were glycosylated. 4. Proteins migrating similar to these three Novikoff hepatoma fractions were detected by polyacrylamide gel electrophoresis in the chromosomal nonhistone protein fraction obtained from normal rat liver. 5. However, the three proteins from normal liver were PAS negative, and did not react immunologically with antiserum to dehistonized Novikoff hepatoma chromatin.

Animals↗
PubMed1982

Identification of cytokeratin antigens in Novikoff ascites hepatoma.

Further characterization of three polypeptide antigens present in Novikoff ascites hepatoma (molecular weights 39,000, 49,000, and 56,000--p39, p49, and p56, respectively), has established the identity of these proteins as cytokeratins. Two-dimensional gel electrophoresis revealed that each of the three proteins exists in several isoelectric forms, all characteristic of the keratins. The antigens p39, and p56 had amino acid compositions compatible with this family of proteins, and immunofluorescence microscopy localized the antigens on elaborate filament arrays indicative of the cytokeratins. Antibodies monospecific for the p39 antigen, purified by using affinity chromatography, were used to demonstrate that protein p39 was specific for Novikoff hepatoma cells. These results suggest that the cytokeratins contain cell-specific that can be useful as markers for differentiation and neoplasia.

Amino Acids↗
PubMed1982

Antigenic changes in nonhistone proteins during azo dye hepatocarcinogenesis.

We have investigated the appearance of specific nonhistone proteins during azo dye-induced hepatocarcinogenesis in the rat. Groups of animals fed azo dye-containing diet were sacrificed at approximately 3-week intervals, portions of their livers were examined histologically, and the remaining material was fractionated into chromatin and cytoplasmic fractions. Livers of the azo dye-fed animals exhibited histological changes that have been classically attributed to the course and development of cancer; by 28 to 30 weeks of treatment, nearly all animals had developed hepatomas. Heterogeneous rabbit antisera were prepared to dehistonized chromatin from several azo dye-induced hepatomas. These antisera were then used to assess various chromatins for the appearance of antigens specific for neoplasia during inducing carcinogenesis using immunodetection of antigens separated electrophoretically and transferred to nitrocellulose. Changes in the immunoreactivity of liver chromosomal proteins during carcinogen treatment were evident after 3 weeks, and the antigenic profiles of various chromatin samples gradually assumed the characteristics of the hepatoma. The transformation was accompanied by qualitative changes in chromosomal protein antigens, and although these antigenic species were not directly quantitated, noticeable enrichment of tumor-specific species occurred with treatment time. Immunotransfer assays of cytoplasmic fractions indicated most antigens to be specific for chromatin. Normal tissue chromatin exhibited minimal immunoreactivity, and slightly more antigenic homology was noted with regenerating liver and most transplantable tumor chromatins. Interestingly, the transplantable tumor Walker 256 carcinosarcoma was highly enriched in antigens recognized by antisera to azo dye hepatoma dehistonized chromatin. These studies establish a define chronological correlation between the chemical induction of cancer and sequential changes in the immunological specificity of nonhistone protein antigens.

Animals↗
PubMed1982

Tumor-associated chromatin antigens of human colon adenocarcinoma cell lines HT-29 and LoVo.

Antiserum to dehistonized chromatin from human colon adenocarcinoma cell line HT-29 was raised in rabbits and used for the immunodetection of colon tumor-associated nuclear antigens. Quantitative microcomplement-fixation studies indicated that the antiserum had reactivity similar to those of chromatins of two human colon adenocarcinoma cell lines (HT-29 and LoVo) and with nonneoplastic colon. Immunocytochemical localization showed the chromatin antigens to be evenly dispersed throughout the nucleus. Immunochemical staining of electrophoretically separated chromosomal proteins transferred to nitrocellulose sheets revealed several intensely staining antigens predominantly in the molecular weight range of 62,000 to 75,000. When the antiserum was immunoabsorbed with nonneoplastic human colon chromatin, two antigens with molecular weights of 67,000 and 92,000 were identified only with the tumor chromatins. Antiserum immunoabsorbed with either LoVo or HT-29 chromatin failed to show any immunoreactivity, suggesting that these tumor-associated proteins, or antigenic sites residing on these proteins, are identical in both colon tumor cell lines.

Adenocarcinoma↗
PubMed1982

A nuclear matrix antigen HeLa and other human malignant cells.

Antisera were obtained in rabbits to preparations of dehistonized chromatin from HeLa cells. By complement fixation assays, the antisera reacted with HeLa cell chromatin but only marginally with human placenta chromatin. The complement-fixing reactivity of the antisera was inversely related to the amount of dehistonized chromatin used for immunization. Immunochemical staining of electrophoretically separated chromosomal proteins transferred to nitrocellulose sheets revealed numerous antigens in chromatin preparations from several human tumors, placenta, and normal kidney. While immunoabsorption of the antisera with placenta chromatin removed some of the immunochemical staining, many of the electrophoretically separated antigens resisted repeated immunoabsorptions. However, further comparisons revealed that only one major protein antigen (band at an approximate molecular weight of 81,000) was represented in all the assayed human tumors while being absent from human placenta or kidney. Fractionation of HeLa cells into three cytoplasmic and several nuclear fractions showed that almost all the antigens recognized by antisera to dehistonized chromatin were nuclear. The antigenic protein with an approximately molecular weight of 81,000 was found associated with the nuclear matrix fraction.

Antigens↗
PubMed1981

Tissue specificity and cellular distribution of Novikoff hepatoma antigenic proteins p39, p49, and p56.

Three Novikoff hepatoma protein antigens (approximately Mr = 39,000, 49,000, and 56,000) were partially purified from Novikoff hepatoma chromatin. Rabbit antiserum to these three proteins was used to examine various rat tissues for the presence of these antigens. Immunological specificity of the antiserum was assessed using quantitative microcomplement fixation assay or by visualization of the immunoreactive complexes with peroxidase-antiperoxidase procedure after transferring the electrophoretically separated proteins to nitrocellulose sheets. The immunoreactivity was localized with the three proteins p39, p49, and p56 in Novikoff hepatoma. The p56 protein was found to be present in normal rat liver, 24-h regenerating rat liver, fetal rat liver, or kidney, albeit in much smaller amounts as found in Novikoff hepatoma. the p49 and p39 antigens were specific for Novikoff hepatoma. Immunoabsorption experiments confirmed the specificity of this antiserum. Assessment of various subcellular fractions of Novikoff hepatoma revealed that the p39, p49, and p56 protein antigens are present in the cytoplasmic fractions as well as in isolated chromatin.

Animals↗