The use of brominated toluidine blue O in X-ray microanalysis for proteoglycan

Toluidine blue O, a cationic dye has been used for visualizing proteoglycan by light and electron microscopy. Toluidine blue O has now been brominated and incorporated into epiphyseal cartilage during its fixation. Using energy dispersive X-ray microanalysis, bromine has been measured from area to area and compared to measurements of sulphur, a naturally occurring element of proteoglycan. Since bromine has been combined mole for mole with toluidine blue O and it is already known that toluidine blue O combines approximately mole for mole with the glycosaminoglycans of proteoglycan, the measurement of bromine, presents an indirect measure of proteoglycan concentration. The X-ray spectral localization of sulphur presents problems in X-ray analysis which are overcome with bromine which makes this technique attractive for workers in this field.