The size of bacteria, as measured by interference microscopy

SYNOPSIS Bacteria were examined with a Smith interference microscope using a half‐shade eyepiece and photomicrographs of their images were prepared. Microdensitometer traces on the negatives demonstrated conclusively that the method developed by Ross in 1957 to measure the thickness of bacteria was subject to systematic error. This error was eliminated by modifying the half‐shade eyepiece so that its comparison area was no larger than the image of a single bacterium; a critical comparison of this new method of measurement of phase change through small microscopic objects demonstrated its advantages over the other well‐established methods. Measurements of bacterial thickness obtained by this new method are markedly lower than those obtained by other interferometric methods; they are, however, still slightly higher (by a maximum of 30 p.c.) than the values obtained by measuring the widths of similar bacteria directly from the mid‐points of the diffraction fringes visible at the boundaries of the image. The work of Wolter and of Charman is discussed in the light of these findings.