1. ADP-dependent platelet aggregation is mediated by the P2T-purinoceptor and is specifically inhibited by ATP, which is a competitive P2T-purinoceptor antagonist. However, ATP functions as an agonist at other P2-purinoceptor subtypes in other tissues and is, therefore, non-selective. This paper describes the effects of the novel ATP analogue, FPL 66096 (2-propylthio-D-beta,gamma-difluoromethylene ATP), on ADP-induced and ADP-independent aggregation of human washed platelets and in standard preparations containing P2X- (rabbit ear artery) and P2Y-purinoceptors (guinea-pig aorta). 2. In suspensions of human washed platelets, FPL 66096 (1-100 nM) produced concentration-dependent rightward displacement of concentration-effect (E/[A]) curves obtained for ADP-induced platelet aggregation. Logistic fitting of E/[A] data indicated that the effect of FPL 66096 was consistent with simple competition with a pKB value of 8.66. FPL 66096 (10-1000 nM) had no effect on aggregation produced by the thromboxane A2-mimetic, U46619 (0.1-10 microM) when the response to this agent was rendered ADP-independent by inclusion of the non-selective P2-purinoceptor antagonist, suramin (100 microM). 3. The anti-aggregatory potency of FPL 66096 was not influenced by increasing the incubation time from 2 to 15 min nor by inclusion of the P1-purinoceptor antagonist 8-sulphophenyltheophylline at a concentration (300 microM) that produced a 68 fold rightward displacement of the anti-aggregatory E/[A] curve for the P1-purinoceptor agonist, 5'-N-ethylcarboxamidoadenosine (0.1-1000 microM). 4. FLP 66096 behaved as a weak (pA" 3.68) but full P2x-purinoceptor agonist in preparations of the rabbit isolated ear artery and as a weak, competitive antagonist (apparent pKB 4.71) at P2Y purinoceptors in the guinea-pig isolated aorta, indicating a selectivity of at least 9000 fold for the P2t-subtype. In the latter preparation, non-specific relaxations were produced by concentrations of FPL 66096 >10M gM.5. These results indicate that FPL 66096 is a P2-purinoceptor antagonist of unprecedented potency and selectivity and that its effects are consistent with simple competition at the P2-purinoceptor. Therefore,FPL 66096 represents a novel pharmacological tool in the classification of P2-purinoceptors and in the elucidation of the mechanisms involved in activation of platelets by ADP.