Objectives: Non-painful needle sensations elicit different hemodynamic responses compared to painful needle sensations which may indicate positive therapeutic benefits. Yet there is evidence regarding the perception of painful and non-painful sensations during acupuncture. This study measured painful and non-painful sensations during and after the insertion of a single acupuncture needle. Design: This study used a repeated measures, crossover design. Setting: This experiment was conducted in the physiology laboratory at Leeds Metropolitan University, UK. Subjects: Fifteen healthy participants were recruited from university staff and students. Intervention: During each session, a single acupuncture needle was inserted at LI10 for 15 minutes, and needle sensations were recorded at 5-minute intervals for a total of 30 minutes. During the first visit, an acupuncture needle was inserted to a depth of 15-25 mm followed by bidirectional rotation (Experimental condition). During the second visit, an acupuncture needle was inserted to a depth of 5mm with mock bidirectional rotation (Control condition). Main Outcome Measures: Needle sensation intensity was measured using a visual analog scale. Painful and non-painful characteristics of needle sensation were reported on the Massachusetts General Hospital Acupuncture Sensation Scale. Participants were also asked to report if needle sensations were painful or not by indicating yes or no. Results: Bidirectional needle rotation increased the overall needle sensation intensity and the intensity of painful and non-painful sensations. Nine participants reported sensations as painful during bidirectional rotation of the needle and no participants reported painful sensations during mock rotation of the needle. There were no differences in sensation characteristics between the participants reporting painful or non-painful sensations except that of numbness which was lower in participants reporting painful sensations compared with those that did not. Conclusions: Bidirectional rotation of an acupuncture needle inserted at a depth of 15-25mm produced stronger painful and non-painful needle sensations compared to mock bidirectional rotation of a needle inserted 5mm into the skin.