RESUMEN
Lipoma cells with consistent chromosomal aberration have been transfected with plasmids carrying papilloma bovine virus subgenomic fragment (PBV 69). The successful transformation of the cells was ascerted on the changed growth pattern of the cells in liquid medium, colony formation in soft agar and modified cell appearance in electron microscopy; transfection with PBV 69 has not been, however, sufficient to immortalize lipoma cells.
Asunto(s)
Papillomavirus Bovino 1/genética , Transformación Celular Viral , Lipoma/patología , Transfección , Tejido Adiposo/patología , Diferenciación Celular , División Celular , Línea Celular Transformada , Aberraciones Cromosómicas , Células Clonales , Retículo Endoplásmico/ultraestructura , Fibroblastos , Humanos , Lipoma/genética , Microscopía Electrónica , Músculos/patología , Células Tumorales CultivadasRESUMEN
In this paper we present a simple colorimetric method for evaluating cell growth in adhering cell cultures. This technique is based on the staining of basophilic cellular compounds (mainly nucleic acids) with methylene blue. To check its reliability, we have compared the quantity of dye fixed by the cells with the number of cells released from the culture substratum by trypsinization, and with the total cellular protein content determined by the Lowry's method. In these experiments we have used human skin fibroblasts and human epithelial cells derived from the epithelium of the full-term umbilical cord. Like other alternative methods, the methylene blue colorimetric technique can be used in 96-well microplates and allows the rapid collection of large amounts of data. As an illustration, we report on the selection of optimal composition of culture medium for human skin fibroblasts.
Asunto(s)
Colorimetría , Fibroblastos/citología , Adhesión Celular , División Celular , Células Cultivadas , Medios de Cultivo , Células Epiteliales , Humanos , Cinética , Azul de Metileno , Coloración y Etiquetado , Azul de Tripano , Cordón UmbilicalRESUMEN
Repeated trypsinization of the full term human umbilical cord epithelium allows an homogeneous and exclusively epithelial primary culture, without fibroblastic growth. Transmission electron microscopy observations of desmosomes, cytokeratin intermediate filaments as revealed by indirect immunofluorescence and cultural aspects confirm the epithelial nature of this primary culture. Fibroblasts obtained by an explants culture method exhibit neither desmosomes nor cytokeratin intermediate filaments, which are epithelial markers. They yield characteristic long term cultures of fibroblastic aspect and growth.
Asunto(s)
Cordón Umbilical/citología , Células Cultivadas , Células Epiteliales , Epitelio/ultraestructura , Fibroblastos/citología , Fibroblastos/ultraestructura , Técnica del Anticuerpo Fluorescente , Humanos , Microscopía Electrónica , Cordón Umbilical/ultraestructuraRESUMEN
A short-term method for detecting potential environmental carcinogens is described using in vitro transformation assay of human epithelial-like amniotic fluid cells. According to Styles, after a short exposition to the carcinogen, only transformed cells grow and form large colonies when cultured in semi-solid agar. In our assay addition of liver homogenate was not necessary to activate the carcinogens. Nevertheless adjunction to the exposure medium of human fecalase (after Ames) is advised for studying carcinogenicity of food glycosides. Fecalase efficiency in metabolic activation of glycosidic compounds has been demonstrated in the use of 8-hydroxyquinoline-beta-D-glycopyranoside.