Fresh and cultured buccal cells as a source of mRNA and protein for molecular analysis.
We developed a method for obtaining viable buccal cells from mouthwash samples for use as a source of mRNA and protein. Immunofluorescent analysis showed that most cells were derived from nonkeratinized parabasal epithelia, with a minor proportion of proliferative cells. Gene expression was detected in buccal cells using reverse transcription PCR, Western blot analysis, and immunofluorescence. Using a keratinocyte-specific medium, buccal cells could be cultured on Matrigel-coated permeable filters for up to 2 weeks while maintaining the expression of some epithelial-specific markers, including cytokeratin 13, cytokeratin 10, transferrin receptor, and beta-integrin. The basal marker cytokeratin 14 and Ki67, an indicator of cellular proliferation, were detected in a few cells. We show that buccal cells can be obtained from a noninvasive procedure for use as a source of material for biochemical analyses. A population of the buccal cells can be maintained in culture for up to 2 weeks using keratinocyte-specific medium in combination with extracellular matrix.
Michalczyk
Agnes
A
Deakin University, Melbourne, VIC, Australia.
Varigos
George
G
Smith
Lance
L
Ackland
M Leigh
ML
D018929:Cell Culture Techniques / Q000379:methods*; D049109:Cell Proliferation; D002470:Cell Survival / Q000502:physiology; D005109:Extracellular Matrix / Q000502:physiology*; D006801:Humans; D007633:Keratins / Q000378:metabolism*; D009061:Mouth Mucosa / Q000166:cytology* / Q000502:physiology*; D009067:Mouthwashes; D012333:RNA, Messenger / Q000302:isolation & purification / Q000378:metabolism*; D013048:Specimen Handling / Q000379:methods*
D009067:Mouthwashes; D012333:RNA, Messenger; D007633:Keratins