Cell experiment:

L929 cells in 2 mL DM10F are allowed to adhere on microscope slide flasks. For the observation of plasma membrane labeling, cells are incubated for a short time (10 s) at room temperature with TMA-DPH 2×10-6 M in PBS or in DM10F from a 4×10-3 M stock solution in dimethylformamide. The unwashed slide is then transferred to the microscope and observed. For the labeling of internalized membrane, the cells are incubated in slide flasks at 37℃, with TMA-DPH 2×10-6 M in DM10F for the desired time, and then washed by gently shaking the slide in PBS for a few seconds[1].

TMA-DPH is a fluorescent probe used for measuring membrane fluidity in artificial and living membrane systems.[1],[2],[3],[4] The cylindrical shape of TMA-DPH confers high sensitivity to reorientation resulting from changes in surrounding lipids. TMA-DPH has excitation and emission maxima at 355 and 430 nm, respectively.

Reference:
[1]. Subramanian, V., Knight, J.S., Parelkar, S., et al. Design, synthesis, and biological evaluation of tetrazole analogs of cl-amidine as protein arginine deiminase inhibitors. Journal of Medicinal Chemistry 58(3), 1337-1344 (2015).
[2]. Illinger, D., and Kurhy, J.G. The kinetic aspects of intracellular fluorescence labeling with TMA-DPH support the maturation model for endocytosis in L929 cells. Journal of Cell Biology 125(4), 783-794 (1994).
[3]. Illinger, D., Duportail, G., Mely, Y., et al. A comparison of the fluorescence properties of TMA-DPH as a probe for plasma membrane and for endocytic membrane. Biochimica et Biophysica Acta 1239(1), 58-66 (1995).
[4]. Chazotte, B. Labeling the plasma membrane with TMA-DPH. Cold Spring Harb.Protoc. 2011(5), (2011).).