Table 1.

Summary of the ICS staining buffers used and the quality of RNA obtained

Buffer	Composition	Yield (ng)*,#	280/260 ratio*	RIN*
standard permeabilization buffer	PBS containing FBS (2%), sodium azide (0.05%) saponin (0.1%) and normal rat serum (10%); pH 7.4	575.7 (477.0 – 615.0)	2.01 (1.81 – 2.13)	3.8 (2.3 – 5.2)
RNA preserving hybridization buffer	2× SSC containing ammonium sulfate (2.1 M), EDTA (10 mM), ultrapure BSA (1 mg/ml), formamide (25% [40% v/v]) and saponin (0.1%); pH 5.2	NP	NP	NP
high salt buffer	PBS containing sodium azide (0.05%) saponin (0.1%), purified rat IgG (0.02 mg/ml), ultrapure BSA (1 mg/ml) and sodium chloride (2.0 M); pH 7.4	585.3 (416.4 – 747.0)	2.03 (1.72 – 2.15)	9.1 (7.8 – 10.0)
buffer containing RNase inhibitor	PBS containing sodium azide (0.05%), saponin (0.1%), purified rat IgG (0.02 mg/ml), ultrapure BSA (1 mg/ml) and RNase inhibitor (RNasin® Plus RNase Inhibitor, 0.5 U/ml); pH 7.4	574.9 (492.0 – 645.0)	1.98 (1.70 – 2.13)	9.0 (8.1 – 10.0)

^*average value with range in parenthesis

^#total yield per million cells

NP not performed

Results represent two independent experiments (n = 3)