Polymorphic DNA markers were used to identify eight sublines derived from HeLa. Using five highly polymorphic minisatellite DNA probes, these cell lines were distinguished and classified into six groups by Southern blot analysis. Polymorphic DNA markers, therefore, can provide a useful tool for monitoring genetic changes of a cell line during culture and for distinguishing sublines derived from the same origin.

Blotting, Southern↗

PubMedDate not supplied

Clastogenic potential of heavy oil extracts and some aza-arenes in Chinese hamster cells in culture.

Chromosomal aberration tests in vitro, with a Chinese hamster fibroblast cell line, CHL, were carried out on B class heavy oil fractions, obtained by silica-gel column chromatography and a liquid-liquid extraction method. The original oil and basic nitrogen-containing fractions induced structural chromosomal aberrations in the CHL cells in the presence of rat-liver microsome fraction (S9 mix). 8 tricyclic or pentacyclic aza-arenes, which possibly exist in the positive fractions, were also examined for their clastogenic activities in the system with or without S9 mix. Acridine, benzo[f]quinoline, pyrenoline and pyrenoline 4,5-oxide induced chromosomal aberrations both with and without S9 mix. Benzo[h]quinoline was positive without S9 mix, whereas dibenz[c,h]acridine and dibenz[a,j]acridine were only positive with S9 mix. The results suggest that the clastogenic effects of the heavy oil may in part be due to the presence of nitrogen-containing polycyclic hydrocarbons such as aza-arenes.

Animals↗

PubMedDate not supplied

Micronucleus induction in mouse peripheral reticulocytes by 7,12-dimethylbenz[a]anthracene.

Micronucleus assays using mouse peripheral blood stained vitally on acridine orange (AO)-coated slides were evaluated at two laboratories with 7,12-dimethylbenz[a]anthracene (DMBA) and compared with the standard bone marrow assay. DMBA was administered by single intraperitoneal injection to CD-1 mice at doses ranging from 5 to 80 mg/kg, then 5 microliters of peripheral blood was sampled from a tail vein at 24, 48, 72, 96, and 120 h after treatment. Similar incidences of micronucleated young erythrocytes were observed in peripheral blood reticulocytes and bone marrow polychromatic erythrocytes. The dose response of micronucleated reticulocytes was delayed compared to that of micronucleated polychromatic erythrocytes. The dose-response curves after treatment with DMBA differed depending on the sampling times, which revealed the difficulty of obtaining accurate dose-response relations in the micronucleus assay. The present result demonstrated that the simple and rapid AO supravital staining method is a valuable and easier method for obtaining dose- and time-response data for quantification of micronucleus induction by chemicals.

9,10-Dimethyl-1,2-benzanthracene↗

PubMedDate not supplied

The micronucleus assay using peripheral blood reticulocytes from mitomycin C- and cyclophosphamide-treated rats.

It used to be believed that the use of rat peripheral blood for the micronucleus assay would be difficult because micronucleated erythrocytes are captured and destroyed by the spleen quickly. We have applied an acridine orange (AO) supravital staining method to rat peripheral blood using AO-coated glass slides. Normal and splenectomized SD rats were treated once with mitomycin C (i.p.) or cyclophosphamide (p.o.), and 5 microliters of blood was collected at intervals from the tail vein between 0 and 72 h after treatment. For comparison, bone marrow cells were smeared conventionally 30 h after treatment. Although the frequencies of spontaneous and chemically induced micronucleated reticulocytes (MNRETs) from normal rats were lower on average in the highest dose group than those of splenectomized rats, the incidence of micronuclei among type I and II reticulocytes in normal rats at 48 h was almost identical to the incidence of RNA-containing erythrocytes with micronucleus in bone marrow. Thus, we suggest that rat peripheral reticulocytes can be used as target cells for the micronucleus assay.

Acridine Orange↗

PubMedDate not supplied

A pilot experiment for the micronucleus test. The multi-sampling at multi-dose levels method.

A pilot experiment was undertaken to find the optimal dose range and sampling time in the micronucleus test; 2 animals per group were used. The chemical was injected at 4 different doses, and smear preparations were made from femoral marrow cells at 5 different sampling times for each dose group. The incidence of micronucleated polychromatic erythrocytes was scored on the preparations stained with acridine orange. When the data were arranged two-dimensionally, the optimal dose and sampling time which could be used for a further full-scale test, could be estimated. In addition, sex differences and the effect of multiple treatment can be estimated using this protocol with minor modifications.

Animals↗