Application of immunocytochemistry and immunofluorescence techniques to adipose tissue and cell cultures.
When isolated from tissue, white adipose cells are round, and their interior is filled with a large (80-120 microm) droplet of stored triglyceride, leaving a thin (1-2-microm) layer of cytoplasm between the lipid droplet and the plasma membrane. Their three-dimensional architecture, together with the fact that these cells ordinarily float in medium, have created major challenges when one attempts to perform microscopy techniques with these cells. Adipocytes serve as the principal energy reservoir in the body, and it is essential to overcome these difficulties to be able to study hormone-mediated responses in real adipose cells, which convey physiological significance that cannot be readily duplicated by the use of cultured model adipocytes. This chapter focuses on the use of confocal microscopy optical sectioning and computer-assisted image reconstruction in the whole adipose cell in the study of insulin-regulated protein trafficking. In addition, we illustrate the possibility to image whole-mount preparations of living adipose tissue, opening new ways to probe adipose cells in situ without disrupting their cellular interactions within living adipose tissue. Confocal microscopy constitutes an effective morphological approach to investigating adipose cell physiology and pathophysiology.
Light Microscopy Core Facility, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD, USA.
Daniela
D
Malide
D017667:Adipocytes / Q000166:cytology / Q000378:metabolism; D000273:Adipose Tissue / Q000166:cytology / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D004822:Epididymis / Q000033:anatomy & histology; D005455:Fluorescent Antibody Technique; D051275:Glucose Transporter Type 4 / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007150:Immunohistochemistry / Q000379:methods*; D007328:Insulin / Q000378:metabolism; D008297:Male; D018613:Microscopy, Confocal / Q000295:instrumentation / Q000379:methods; D051381:Rats
D051275:Glucose Transporter Type 4; D007328:Insulin; D049452:Green Fluorescent Proteins