ABSTRACT
This chapter describes some immunohistochemical methods and their application to the study of the pulmonary extracellular matrix (ECM). It discusses some theoretical considerations, then specific protocols, and finally reviews the contributions of immunohistochemistry to the understanding of the pulmonary ECM. The goal of tissue preparation is to stabilize tissue components, which can be achieved by physical or chemical methods. Optimal fixation is usually achieved by cross-linking tissue components, which may result in loss of antigenicity. The goal of postfixation treatments is to quench and remove excess fixative and prepare tissue for the subsequent steps. Most embedding procedures involve the replacement of water, either by the embedding media or by an intermediate solvent. Free aldehyde radicals will react with e-amino groups; therefore, immunoglobulin molecules will bind to aldehyde fixed tissues by a nonimmune mechanism, resulting in high backgrounds. Polyclonal antibodies have the advantage of recognizing multiple epitopes in a molecule and having high affinities.
