Background: We investigated the reproducibility of contrast transcranial Doppler (TCD), a safe non-invasive test for investigation of venous-to-arterial circulation shunts (v-aCS), usually patent foramen ovale, in young stroke patients. We also investigated whether microbubble contrast was reproducible and whether the addition of blood to agitated saline contrast affected the number of microbubbles produced. Methods: TCD investigation for v-aCS was repeated in 42 patients using a standardised protocol (i) by the same investigator and (ii) by a different investigator. Agitated saline was produced by mixing saline and 1 ml of air between two 10-ml syringes. The effect of adding blood and increasing the number of agitations was evaluated by microscopy examination using a haemocytometer to assess bubble numbers and sizes. Results: TCD: no difference was found in the highest microbubble count for the same investigator and between different investigators (p > 0.05). Reproducibility for the detection of v-aCS consistent with a patent foramen ovale was also good (ĸ values >0.8). Contrast: both the number of contrast mixes before injection and the presence of blood significantly increased the number of bubbles counted. On average, 18 agitations produced 1.86 (95% CI 1.62–2.13) times more bubbles than 6 agitations. Mixtures with blood produced on average 3.8 times more bubbles (3.08–4.69). The size of the bubbles was similar for all mixtures. Conclusions: Contrast TCD is reproducible and reliable for the detection of v-aCS. The addition of blood and 18 mixes rather than 6 significantly increased the number of microbubbles produced and may increase the effectiveness of microbubble contrast.

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