14] Measurement of tissue thickness using confocal microscopy

Abstract

Publisher Summary The chapter presents a discussion on measurement of tissue thickness using confocal microscopy. The major technical details concerning the in vivo measurement of tissue thickness using reflected light confocal microscopy are discussed. Critical to the in vivo measurement of tissue thickness using confocal microscopy is the imaging of tissue in real time. This is necessary to avoid unwanted irregular movement of the tissue because of pulse and respiration and to capture fast temporal events, such as blood flow or acute cellular responses, which can blur images at high magnification. There are several advantages to the tandem scanning confocal microscope (TSCM) for use in evaluating living tissue, first of which is the real time imaging capabilities. The white light used is also not toxic to tissues and produces minimal discomfort. Additionally, the use of diffraction-limited pinholes provides optimal lateral and axial resolution when using objectives of the same numerical aperture. As an alternative to the tandem scanning design, a one-sided Nipkow disk system where illumination and detection occur simultaneously through the same pinhole is developed. The chapter also discusses the measurement of tissue thickness by using a depth encoding system (DES) and by rapid through focusing approach.