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In 1974, the presence of intracytoplasmic immunoglobulins in plasma cells was demonstrated in routinely processed paraffin tissue (the mainstay of diagnostic pathology) using an immunoperoxidase technique (1). This new capability has introduced the surgical pathologist to a t ...

A significant consideration in the use of immunoreagents for microscopic slide staining is their cost. For many antibodies, fractions of a milliliter of fluid may be very expensive, particularly in the undiluted form. Traditional staining methods frequently use more reagent than allo ...

Fluorescence in situ hybridization (FISH) expands the repertoire of user-friendly (nonradioactive), signal-generating systems, which may be used to identify and characterize chromosomes, interphase nuclei, and other sources of nucleic acids in biological material. FISH is b ...

Protein A chromatography is a type of affinity chromatography that relies on the specific interaction of protein A with the Fc region of immunoglobulins from a number of species (1). Protein A is a 42,000-Dalton polypeptide originally isolated from the cell walls of Staphylococcus aureus (2). ...

Important aspects of the relatively new technique, nucleic acid immunocytochemistry, are predicated on principles that are different from those that underlie the immunocytochemistry protocols described in earlier chapters of this book. Hence, this chapter includes selected ...

When fluorescently labeled biological specimens are viewed with a conventional wide-field microscope, a haze of out-of-focus fluorescence is usually created by the overlapping structures within the sample. As we focus through the specimen, our brains have a remarkable ability to dis ...

The results of fluorescence-labeling experiments may be photographed to produce a permanent record. One of the major hurdles associated with fluorescence photography is that most of the commercially available films have been designed to work at exposure times in the range of 0.0l– 0.1 s. Howe ...

Since it was first introduced (1), postembedding immunogold labeling has become the most widely used method of immunolabeling for electron microscopy (2–8). For postembedding labeling, samples are first fixed, embedded, and sectioned. All immunostaining is performed on sections m ...

Colloidal gold conjugates generally do not readily penetrate cells, even after permeabilization. Therefore, their use in pre-embedding immunostaining has been restricted to labeling cell-surface antigens for scanning (1) or transmission electron (Fig. 1A) microscopy or for tr ...

Biotin-avidin detection systems are widely used in both immunocytochemistry and molecular biology (1, 2) (see Chapter 21). They take advantage of the high affinity of biotin, a low-molecular-weight vitamin, for avidin, an egg-white protein. The avidin-biotin complex has one of the highest ...

The ability to conjugate proteins to colloidal gold sols provides a wide variety of probes for electron microscopy. In addition to antibodies, protein A, lectins, enzymes, toxins, and other proteins have all been conjugated to colloidal gold (1– 7). The nature of the interaction between the coll ...

Immunocytochemistry, by definition, is the identification of a tissue constituent in situ by means of a specific antigen-antibody interaction where the antibody has been tagged with a visible label (1) Cell staining is a powerful method to demonstrate both the presence and subcellular lo ...

Since their introduction by Faulk and Taylor (1), colloidal gold probes have become widely used for immunocytochemical staining at the electron microscopic level. Many different methods of producing colloidal gold sols have been published (2–10). Gold sols are producing by boiling a sol ...

For electron microscopic immunocytochemistry, the fixation procedure is always a compromise between good morphological preservation and retention of antigenicity (1, 2). It is preferable to fix tissues by perfusion, but if that is not possible, the time between removal of the tissue and f ...

The effectiveness of affinity chromatography relies on the ability of a molecule in solution to recognize specifically an immobilized ligand (1–3). This type of separation, unlike other chromatographic methods, uses the intrinsic biological activity of a molecule to bind to a substra ...

Phagocytic leukocytes that are exposed to opsonized particles, chemoattractants, or selected cytokines undergo a rapid burst in oxygen consumption and activation of the enzyme responsible for the oxidative metabolic burst, NADPH oxidase (reviewed in 1). Active NADPH oxidase cat ...

Phagocytic leukocytes play a major role in host defense because they rapidly migrate to sites of infection and destroy invading microorganisms. Specific signal molecules (chemoattractants), released by bacteria or endogenously generated by the host, can elicit directed leukoc ...

Phagocytic leukocytes (granulocytes, monocytes, and macrophages) provide a first line of host defense by their ingestion (phagocytosis) and killing of microorganisms. Leukocytes phagocytize particles through a complex series of events that include serum coating (opsoniz ...

DNA content has become an important diagnostic, as well as prognostic, method for clinical pathology and investigative oncology. Paraffin-embedded tissue can be examined by flow cytometric (FCM) methods for total DNA content and aneuploidy with respect to the classification of the or ...

Flow cytometry (FCM) is a high-precision technique for rapid analysis and sorting of cells and particles. In theory, it can be used to measure any cell component, provided that a fluorescent tracer is available that reacts specifically and stoichiometrically with that constituent. The tec ...