much more robust than inhibition with monovalent aptamers . This data suggests that the LS-Multi-Aptamer can inhibit binding to endogenous ligands, either through blocking or inducing 1239358-86-1 shedding of L-selectin from the cell surface. As Naringin previous studies have reported that multivalent polymers and cross-linked antibodies induce L-selectin shedding , we next investigated if the LS-Multi-Aptamer led to shedding of L-selectin. After Jurkat cells were untreated or treated with PMA, LS-Multi-Aptamer, and SC-Multi-Aptamer, we collected the supernatants for analysis by ELISA to detect soluble Lselectin and lysed the treated cells for analysis of cell-associated L-selectin by Western blot . PMA, a well-established inducer of L-selectin shedding, induced L-selectin shedding while the untreated, LS-Multi-Aptamer, and SC-Multi-Aptamer treated cells retained their cell surface L-selectin. Our results therefore suggest that the LS-Multi-Aptamer does not induce shedding of L-selectin but rather blocks its functional binding to endogenous ligands . Although previous studies have reported that multivalent L-selectin ligands induced shedding, shedding induced by synthetic polyvalent L-selectin ligands was modest in comparison to PMA treatment , which raises the question of if L-selectin shedding is necessary for modulation of function. It should also be noted that studies of L-selectin monovalent and bivalent aptamers have not demonstrated shedding . Interestingly, synthetic multivalent carbohydrate ligands only induced shedding of L-selectin at millimolar concentrations while blocking L-selectin function at micromolar concentrations . It has been hypothesized that shedding of L-selectin by multivalent ligands depends on both the number of ligands as well as the distance between the ligands . In the future, we plan to tailor the parameters of the LS-Multi-Aptamer to directly determine if the physical properties of the Multi-Aptamers can be optimized to mediate shedding versus blocking. For example, the length of the Multi- Aptamer can be adjusted by varying the duration of the RCA reaction and the distance between aptamers can be modified by adjusting the length of the poly spac
