Figure 2.

Determination of bead positions in the optical trap during pulling DRG neuron membrane tethers. (A) Optical artifacts and their cancelation for an optical trapping experiment. Optical artifacts are best demonstrated in the attempt that does not pull out a membrane tether. (Left) A trace shows optical artifacts (downward deflections) caused by DRG cell shadow when the cell is moved forward and backward near the optically trapped bead. The artifacts due to the forward and backward movement are symmetric. (Right) Artifact cancelation is made by subtracting the mirror points of the trace with the alignment at the two points indicated by the open and solid arrowheads. The subtracted values (black trace) yield a straight line at the position of 0 V position. Note that the bead position is registered in the values of voltages by the quadrant photodiode. (B) (Left) A trace shows upward deflection caused by the formation, extension, and holding of a DRG membrane tether. Note that the optical artifacts of the two downward deflections are present before pulling out the membrane tether. (Right) Mirror subtraction of forward trace (gray, upper) from pulling trace (gray, lower) yields the actual bead position (black) during the tether formation, extension, and holding.