How to dissolve all trans retinoic Acid?

Article Differentiation of the SH-SY5Y Human Neuroblastoma Cell Line

Day 0: Plating Cells for Differentiation

See Figure 1 for the differentiation schedule.

Rinse undifferentiated cells with 1x PBS, aspirate, and then trypsinize using 1-2 ml warmed 1x 0.05% Trypsin-EDTA.

When cells are in trypsin, incubate for approximately 3 min in an incubator.

Quench the trypsin by adding 10 ml Basic Growth Media, rinse the sides of the flask or dish, and gently triturate 1-3 times. Transfer contents to a 15 ml conical tube.

Centrifuge for 2 min at 1,000 x g, and aspirate the media while being careful not to disturb the pellet.

Resuspend pellet in 5 ml Basic Growth Media and triturate 1-3 times.

Count cells using a hemocytometer, then dilute using Basic Growth Media to 50,000 cells/ml.

Plate 2 ml of cells per 35 mm2 dish for a total of 100,000 cells per dish and place back into incubator.

6. Day 1: Change Media (Differentiation Media #1)

Aliquot 50 ml of Differentiation Media #1 (see Table 2)and incubate in a 37 °C water bath.

When media is warmed, allow it to equilibrate in an incubator (37 °C, 5% CO2) for at least one hr to establish a proper pH balance prior to use.

Add Retinoic Acid (RA) (see Table 1) to warmed and equilibrated media immediately before adding media to dishes. Note: Retinoic acid is light sensitive and should be stored in dark bottles at 4 °C

Gently aspirate off old media and discard.

Add 2 ml Differentiation Media #1 with RA per 35 mm2 dish and return to incubator.

Article Optimising parameters for the differentiation of SH-SY5Y cel...

https://www.uni-regensburg.de/Anatomie/Minuth/PDF/Proceedings/2007_Constantinescu.pdf

Praveesh Valissery

Maria Laura Matrella You can try to make 1 mM stocks; that does remain stable. The product specification does say it is soluble upto 20mg/ml but it is sometimes tough to dissolve to 100mM. And anything above a final concentration of 100 uM (in culture) seems to cause preciptation even while you add a fully dissolved clear solution into the media.

Also, Retinoic acid is very sensitive to light and heat in solution. So best not to heat dissolve it.

Arman Rahimmi

Dear Maria,

Are you sure about your main idea? SH-sy5y cell line is not a stem cell line to be differentiated into neurons. they are actually a differentiated cell line.

By the way, retinoic acid is soluble in DMSO in much higher concentrations. Check the purity of your powder. If the remnants of powder within the solution is less than 2-3% of the initial amount, I guess you can ignore it and filter your solution to obtain a clear solution.

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