2.3. Rock-II Inhibition Assay

The assay was done as mentioned in ADP-Glo™ Kinase Assay (SER-THR KINASE SERIES: ROCK2 Kinase assay) (Promega, USA) and Y-27632 [(+)-(R)-trans-4-(1-aminoethyl)-N-(4-pyridyl)cyclohexanecarboxamide dihydrochloride] was used as standard drug; luminescence was recorded using Topotecan, USA, Spark 10 M, multimode microplate reader. A vehicle control for 5% DMSO was used in the assay to check the interference. Standard curve for ROCK-II enzyme was done (Figure 2). Serial dilution and IC₅₀ of the AE was performed in triplicate.

ROCK-II enzyme standard curve. x-axis represents the concentration from (40 ng/ml, 20 ng/ml, 10 ng/ml, 5 ng/ml, 2.5 ng/ml, 1.25 ng/ml and 0.625 ng/ml) and Y-axis represents ΔRLU.