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Biomedical subjects

E Stubblefield

Publications and source records attributed to E Stubblefield.

15 recordsLinked to original sources
PubMed1992

Direct formation of microcells from mitotic cells for use in chromosome transfer.

Microcells, cytoplasmic fragments that contain micronuclei composed of one or a few chromosomes, can be generated directly from mitotic cells. Cytochalasin B, which causes nuclear extrusion in interphase cells, has a similar effect on the chromosomes of colcemid-blocked mitotic cells. The forces generated during centrifugation in a Percoll gradient are sufficient to separate the extruded microcells from the parent cell. The chromosomes contained in an extruded microcell form micronuclei during the process, and in all respects are comparable to microcells generated from micronucleated cells except that they are uniformly in the G1 phase of the cell replication cycle. The procedure is probably applicable to all mammalian cells that grow in culture and can be employed to make microcells for the transfer of both intact and fragmented chromosomes.

Animals↗
PubMed1979

Gene localization by chromosome fractionation: globin genes are on at least two chromosomes and three estrogen-inducible genes are on three chromosomes.

Chicken metaphase chromosomes were partially purified by rate zonal centrifugation, and DNA was prepared from each of the fractions of the sucrose gradient. The DNA was digested with various restriction enzymes and subjected to electrophoresis in agarose gels. The DNA was transferred to nitrocellulose filters (as described by Southern), and the filters were hybridized with cDNA probes. Four globin genes alpha A, alpha D, beta, and rho or epsilon are located on at least two chromosomes, and three of the estrogen-inducible genes of the hen oviduct--ovalbumin, ovomucoid, and transferrin--are on three different chromosomes. These experiments also confirm our earlier assignment of the endogenous viral sequence related to Rous-associated virus-0 to a separate (and larger) chromosome than the cellular sequence related to the transforming gene of avian sarcoma virus (cellular sarc), although it now appears that cellular sarc is on a small macrochromosome, rather than on a microchromosome.

Animals↗
PubMed1979

Ultrastructure of double minutes from a human tumor cell line.

Double minutes (dm) have been isolated from human tumor cells by zonal centrifugation and by differential pelleting of chromosome suspsension. These methods allowed collection of dm in sufficient quantity and purity for visualization with electron microscopy. Ultrastructurally, the chromatin fibers in dm resemble thrance fragments was found. When the two isolation protocols were compared, differential pelleting was shown to increase purity twofold to 85% dm by mass. The differential pelleting procedure enables easy collection of dm in sufficient quantity and purity for chemical analysis.

Carcinoma↗
PubMed1976

Correlation of in vivo malignancy with in vitro properties of human-mouse hybrid cells.

The in vivo tumorigenicity of malignant mouse-nonmalignant human somatic cell hybrids was correlated with the in vitro characteristics. The renal adenocarcinoma mouse cell line RAG and the normal, diploid human cell line Wl-38 were used as the fusion parents. Five independent RAG Wl 38 hybrid clones were tested for concanavalin A (Con A) agglutination patterns, in vitro invasiveness, and tumor formation in immunosuppressed mice. Tests on the parental lines showed that RAG cells agglutinated at much lower levels of Con A than the Wl-38 cells. RAG cells overgrew Wl-38 cells in the in vitro invasiveness assay. RAG cells readily formed tumors in untreated adult or young immunosuppressed mice, whereas the Wl-38 cells did not. The five hybrid clones were all similar, since they had Con A agglutination levels intermediate to those of both parents, though patterns were more "tumor-like", overgrew the Wl-38 cells in the invasiveness assay, and formed tumors in immunosuppressed mice.

Adenocarcinoma↗
PubMed1975

Analysis of the replication pattern of Chinese hamster chromosomes using 5-bromodeoxyuridine suppression of 33258 Hoechst fluorescence.

Chinese hamster fibroblasts were synchronized and given 5-bromodeoxyuridine for DNA synthesis except during one hour of the S phase when thymidine was present in the medium. In the next mitosis, chromosomes stained with 33258 Hoechst were banded in appearance when photographed by fluorescence microscopy. The bright regions corresponded to the chromosome segments replicated during the thymidine exposure in the S phase. The segments replicated together during any one hour produced three distinct patterns which were characteristic of early, middle, and late S phase. Most of the fluorescent regions corresponded in size and position with G-bands of these chromosomes. There was no correlation between the staining behavior of a band in G-band procedure and its time-or-replication, i.e., both light and dark G-bands were replicated during early, middle, and late S phase. However, it appears that all of the DNA within a single band is replicated together within one third of the S phase.

Animals↗