TABLE I.

The conventional protocol was performed on three disodium EDTA and sodium heparin anticoagulated human whole blood. Using a t-test assuming unequal variance, the results of average concentration, average RIN, and average C_q value for three samples were compared between disodium EDTA and sodium heparin anticoagulants. There was no significant difference (p > 0.05) between any of the aforementioned values between the two anticoagulants. The volume of the reagents in the conventional protocol was scaled down by a factor of 22, and a separate experiment mixed the two wash buffers 1:1 to reduce the number of wash steps. Using the conventional protocol as a reference group, all measured values of the adjusted protocols were compared to that of the reference group and were not statistically significant. Note: For the “1/22 scale down” concentration values in the elution buffer—This is not the actual yield of the 1/22 scale down protocol, but for comparison purposes, the measured value was multiplied by 22.

Anticoagulant	Protocol	Average RNA yield (ng/μl elution buffer) ± standard deviation	Average actual yield (ng/μl blood)	Expected yield (ng/μl blood)	Average RIN ± standard deviation	Average Cq value ± standard deviation
Disodium EDTA	Conventional protocol	25 ± 7.21	2.5		4.3 ± 3.0	30.06 ± 4.43
	1/22 scale down	54.05 ± 24.42	0.54		3.4 ± 0.5	29.60 ± 0.85
	1:1 Wash buffer mix	26.22 ± 4.00	2.6	1.6–8	4.7 ± 1.9	26.49 ± 1.16
Sodium heparin	Conventional protocol	20 ± 5.67	2		6.7 ± 1.6	29.01 ± 2.97
	1/22 scale down	36.66 ± 15.02	0.37		3.6 ± 0.7	30.25 ± 1.54
	1:1 Wash buffer mix	31.83 ± 14.90	3.2		4.6 ± 0.6	26.78 ± 0.95