The "black hole" phenomenon was further investigated with porcine whole blood under pulsatile flow conditions in a straight rigid tube 120 cm long and of 0.95 cm diameter. A modified Aloka 280 commercial scanner with a 7.5 MHz linear array was used to collect the radio frequency (RF) signal of backscattering echoes from the blood inside the tube. The transducer was located downstream from the entrance and parallel to the longitudinal direction of the tube. The experimental results showed that higher hematocrits enhanced the black hole phenomenon, leading to a more apparent and larger diameter black hole. The black hole was not apparent at hematocrits below 23%. The highest hematocrit used in the experiment was 60%. Beat rates of 20, 40 and 60 beats per minute (bpm) were used, and the black hole became weaker in amplitude and smaller in diameter when the peak flow velocity was increased at each beat rate. These results are consistent with the suggestion in previous work that the black hole arises from insufficient aggregation of red blood cells (RBCs) at the center of the tube because of the low shear rate. At 20 and 40 bpm, the peak flow velocity ranges were 10 approximately 25 cm/s and 18 approximately 27 cm/s, respectively. The black hole was very clear at the minimal peak flow velocity but almost disappeared at the maximal velocities for each beat rate. At 60 bpm, experiments were only performed at one peak flow velocity of 31 cm/s and the black hole was clear. The results showed that the black hole was more pronounced at higher beat rates when the peak velocity was the same. This phenomenon cannot be explained by previous hypotheses. Acceleration seems to be the only flow parameter that varies at different beat rates when peak velocities are the same. Therefore, the influence of acceleration on the structural organization and orientation of RBC rouleaux might be another factor involved in the formation of the black hole in addition to the shear rate. As the entrance length was changed from 110 to 15 diameters (D) in seven steps at the hematocrit of 60%, it was found that a position farther downstream yielded a black hole with a greater contrast relative to the surrounding region, while the backscattering power at the central hypoechoic zone did not increase with increasing entrance length.