Objective To study the anti-platelet aggregation activity of Dendrobium denneanum. Methods A total of 21 rats were randomly divided into the extract 1 group, the extract 2 group, the extract 3 group, the extract 4 group, the extract 5 group, the extract 6 group and the positive control group, with 3 in each group. While 9 guinea pigs were randomly divided into the extract 5 group, the extract 6 group and the positive control group, 3 guinea pigs per group. The platelet aggregation was induced by adenosine triphosphate (ADP) in vitro in rats and arachidonic acid (AA) in vitro in guinea pigs. Plate let rich plasma (PRP) was collected. Each group of PRP was added with different concentrations of Dendrobium denneanum extract 1, 2, 3, 4, 5, 6 or aspirin to intervene. The maximal aggregation rate were detected in 5 min, and to calculate the inhibition rate of platelet aggregation and half of each test substance inhibitory concentration (IC50). Asp was detected in 5 min. The above steps were repeated three times. The inhibition rate of platelet aggregation and the IC50 of each subject were calculated. The maximum aggregation rate of the drug delivery group and the blank control group was compared to evaluate the antithrombotic activity of the Dendrobium with two different induction methods. Results Compared with the blank control group, the maximal aggregation rate induced by ADP of the extract 1 2.399 mmol/L dose group, the extract 2 8.96 mmol/L dose group, the extract 3 17.505 mmol/L dose group , the extract 5 7.836, 3.918 mmol/L dose group, the extract 6 0.755, 0.377 mmol/L dose group (32.57% ± 0.28% vs. 45.20% ± 2.64%), (16.66% ± 3.67% vs. 39.37% ± 8.05%), (0.00% ± 0.00% vs. 32.52% ± 3.05%), (1.11% ± 1.93%, 7.49% ± 9.47% vs. 43.54% ± 10.68%), (8.66% ± 7.50%, 22.71% ± 8.24% vs. 40.65% ± 6.61%) significantly decreased (P<0.05 or P<0.01). Compared with the blank control group, the maximal aggregation rate induced by AA of the extract 5 7.836 mmol/L dose group, the extract 6 0.377 mmol/L dose group (0.00% ± 0.00% vs. 52.97% ± 5.49%), (0.00% ± 0.00% vs. 53.47% ± 6.04%) significantly decreased (P<0.05 or P<0.01). The IC50 of extracts 5 and 6 were 3.143 and 0.100 mmol/L, respectively. Conclusions There were many kinds of ingredients in Dendrobium denneanum which had anti-platelet aggregation activity. Dendrobium denneanum had positive effects on thrombotic diseases. Key words: Caulis dendrobii; Platelet aggregation; Guinea pigs; Rats