Table 2.

Necessary impurity level to cause 20% bias on K_t estimates from transport studies.

Potency of substrate	Kt/Kia	Necessary Xi level to cause bias in Kt
Strong (Kt = 5 μM)	≤ 1	Never a concern when Xi ≤ 10%
	10	≥ 2.5%
	100	≥ 0.5%
Moderate (Kt = 50 μM)	≤ 1	Never a concern when Xi ≤ 10%
	10	≥ 2.5%
	100	≥ 0.25%
	1000	≥ 0.1%
Weak (Kt = 500 μM)	≤ 1	Never a concern when Xi ≤ 10%
	10	> 2.5%
	100	≥ 0.25%
	1000	Trace
	10000	Trace

^aK_t/K_i reflects the ratio of the “true” affinity constant of the substrate of interest versus the inhibition constant of its impurity.