A new simple method of staining exogenous surfactant in experimental research

Two commonly used techniques in experimental lung research have helped to determine which variables influence surfactant distribution within the lung: radioactive labelling of surfactant components and admixture of coloured microspheres to surfactant. However, neither technique allows the description of surfactant distribution at the alveolar level. The aim of this study was to establish a new technique using histology colourants for admixture to exogenous surfactant to make exogenous surfactant visible by light microscopy. In a step by step approach the authors evaluated the properties of a variety of green colourants when added to a natural porcine surfactant preparation for their ability to homogeneously mix with surfactant, to bind to surfactant, to adhere to a glass slide, to not be “overstained” by standard haematoxylin‐eosin and Elastica van Giesson staining, to not influence in vitro surface tension properties of surfactant using a Wilhelmy balance, to not influence oxygenation and ventilation in a lung‐lavage rat model and to preserve their colour and adherence to exogenous surfactant on lung specimens visualized by light microscopy. Only one of the tested green histology colourants (Green Dye®) fulfilled all requirements and showed a brilliant green colour in a distribution pattern typical of surfactant at the alveolar level. It is concluded that the authors have established a new, simple and inexpensive method of staining exogenous surfactant for evaluation of its distribution by light microscopy at the alveolar level.