Microarrays tissue culture cells

Recent comparison of gene expression profiles of von Hippel-lindau-i- versus von Hippel-lindau-ceU lines obtained from the clear cell RCC subtype identified proteins that might serve as candidate molecular markers. Inactivation of VHL is a hallmark in most sporadic clear cell RCC and it occurs early in renal carcinogenesis (Skates and Iliopoulus., 2004 Latifet al, 1993). Furthermore, the profile of secreted proteins could be directly AQ4 determined by analyzing comparatively the genomic and proteomic patterns of these cell lines as well as their conditioned tissue culture supernatants (Ferguson et al, 2004). A combination of cDNA microarray and proteome analyses appears reasonable since not every difference at the transcriptome level will translate into differences at the protein level, whereas posttranscriptional/posttranslational modifications were not detectable by tran-scriptomics. 

The technological challenges for pharmacoproteomics are exceptional (see Table 13.1-3). Protein microarrays are an emerging class of nanotechnology for tracking many different proteins simultaneously. However, translation into the medical practice is very slow. On the other hand, proteomic changes in cultured cell lines might not fully reflect pharmacodynamic interactions because of the lack of the tissue microenvironment [11]. 

Moskaluk, C. A. and M. H. Stoler. 2002. Agarose mold embedding of cultured cells for tissue microarrays. Diagn Mol Pathol 11 234-8. 

We have used the coUagen-glycosaminoglycan (coUagen-GAG) mesh scaffold to probe molecular level cell-biomaterials interactions [11]. We seeded IMR-90 human fibroblasts onto three-dimensional (3-D) collagen-GAG meshes and control surfaces of tissue culture polystyrene (TCPS). Nucleic acids (mRNA) from cells from each culture were isolated, amplified, and hybridized to human genome microarrays (U133A Gene Chip, Affymetrix, Santa Clara, CA). 

The methods described herein encompass (1) preparation of a whole cell lysate from either cell culture or tissue samples, (2) protein lysate microarray printing, (3) immunostaining, and (4) microarray spot analysis. 

Microelectrode arrays are proving to be useful platforms on which collections of fully functioning cells can be grown and manipulated. Moreover, there is increasing evidence that these cell cultures react to chemical stimulation in a similar way to their parent tissue in fully functional animals. Therefore, it is likely that these microarray methods will play increasingly important roles in determining the efficacy, toxicity, and therapeutic levels for new drug treatments. 

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