Tissue Staining

6 Key excerpts on "Tissue Staining"

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  eBook - PDF

  Biomedical Technology and Devices

  • James E. Moore Jr, Duncan J. Maitland(Authors)
  • 2013(Publication Date)
  • CRC Press

    (Publisher)

  A biological stain has been defined as “a dye for making biological objects more clearly visible than they would be unstained” (Lillie, 1969)� Initially, all dyes were of natural origin, obtained by extraction from plants such as crocus (saffron) (Leeuwenhoek, 1791) cited in Baker (1958b), the tree Haematoxylon campechianum (hematoxylin) (Waldeyer, 1863), and the cochineal beetle (carmine) (Goppert and Cohn, 1849), the latter being the first systematically to study dyed tissues with the microscope although, as men-tioned above, von Leeuwenhoek certainly employed dyes� The early history of histological staining has been briefly reviewed by Lillie (1969) and Baker (1958b) and in more detail by, for instance, Lewis (1942)� FIGURE 13.4 ( See color insert. ) An example of a plastic-embedded, microground section of a coronary artery sec-tion stained with hematoxylin and eosin/phyloxine B and containing a stent strut� Notice the clean interface between the plastic and the metal strut� Multinucleated giant cells surround the strut, while neurovascular buds, scattered macrophages/lymphocytes, and fibrous connective tissue can be seen adjacent to the strut� (Scale bar 100 μ m�)

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  eBook - ePub

  Biological Microtechnique

  • Jeremy Sanderson, Mr Jeremy Sanderson(Authors)
  • 2020(Publication Date)
  • Garland Science

    (Publisher)

  6 Staining and Dyeing

  The visibility of the tissue is commonly increased by using compounds which differentially absorb visible light to produce a coloured preparation. Dyes absorb light; a dye appears red because it absorbs all the colours of the spectrum except red light, which is allowed to pass.

  To study internal structural elements (rather than merely to introduce contrast for studying external features) the stain(s) must act selectively so that specific features of the preparation can be studied alongside differently coloured tissues. The most effective results arise from using a basic and acid stain, preferably of complementary colours, in succession. The basic stains, which are generally used for chromatin, are chosen from the ends of the visible spectrum, while the background counter-stains are chosen from the more central region. This is because the eye perceives yellow and green-yellow hues as unsaturated; if small inclusions and organelles (e.g. chromosomes) were to be stained thus, they would effectively disappear against the more highly saturated red or blue background.

  6.1 Nomenclature

  The term ‘staining’ is generally used to include all methods of colouring tissue. Baker (1958) distinguishes between dyeing and staining; the former process involves the tissue taking up only the dye molecule from solution, leaving the solvent. In the latter case both dye solute and solvent are taken up. The simplest distinction between staining and dyeing is that defined by Kiernan (1990) and Boon and Drijver (1985): where colouring is caused by linking a dye molecule and tissue substrate, the process is called dyeing; if the tissue is stained by solution contact alone, the process is called staining. In this book, the term ‘staining’ will be used throughout to avoid confusion. For a complete discussion of the mechanisms of staining and dyeing see Kiernan (1990) and Boon and Drijver (1985). For a more advanced treatise, see Horobin (1982, 1988).

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  Histochemistry

  An Explanatory Outline of Histochemistry and Biophysical Staining

  • Richard W. Horobin(Author)
  • 2014(Publication Date)
  • Butterworth-Heinemann

    (Publisher)

  1.1 What information does biological staining provide? The essence of the information provided is: what/where. Typically we simultane-ously obtain data regarding what is present and also where in the organism/tissue/cell/ organelle it is found. Such information is obtained by first staining the biological material, then viewing the stained preparation using the light or electron microscope. There is considerable variation as to precisely what is stained, we may stain everything, more or less, or stain selectively. When everything stains information may be obtained from observations of the shapes of the various entities. Such essentially morphological data is generated by systems such as the light microscopists classic H & E oversight stain, by the electron microscopists lead citrate and by the biochemists negative staining of macromolecules and virus particles. 1 Selective staining can be used to provide information of diverse kinds. Thus the entities colour-coded, or otherwise selectively visually labelled, may be defined biologically. Examples include tissue elements such as muscle and connective tissue fibres; micro-organisms such as bacteria, fungi and viruses; and sub-cellular structures such as plant cell-walls or the Golgi apparatus . . . the list could be extended indefinitely. When staining such entities the chemical nature of the substrate is informationally irrelevant, indeed may often not be known. Contrast this with methods which selectively stain defined chemical functions, say phospholipids, Fe(III) ions, or the enzyme lactate dehydrogenase. It is instructive to pause here and compare such histochemical demonstrations with standard biochemical assays. The two procedures are complimentary. Thus a his-tochemical method for demonstrating lactate dehydrogenase will commonly yield precise information on the distribution of this enzyme through tissues and cells, but it is unlikely to yield quantifiable results, through see Section 11.5.

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  Conn's Biological Stains

  A Handbook of Dyes, Stains and Fluorochromes for Use in Biology and Medicine

  • Richard Horobin, John Kiernan, Richard Horobin, John Kiernan(Authors)
  • 2020(Publication Date)
  • CRC Press

    (Publisher)

  A few accounts have emphasized the single set of physicochemical factors operating in all methodologies. In English there are the scholarly books of John Baker (Baker, 1958), various monographs by Horobin (1982, 1988) and Horobin and Bancroft (1998) and Lyon (1991), and more recently a fascinating review by Prento (2001). Such accounts have a characteristic fate: they are widely cited, but their larger message is ignored.

  Basic concepts and terminology

  The purpose of biological staining is to generate information, most often to address the following questions.  

  (i) What is this, that we see in the microscope?

  (ii) Where is this found, within the cell, tissue or organism?

  (iii) How much of this is there, or how many of them are present?

   

  To provide answers a staining method must be sufficiently sensitive to detect the biological target. This often depends upon the amount of material present, and on the precise mode of action of the stain. Other things being equal fluorescent reagents and catalytic staining processes favor high sensitivity. Fluorescent reagents provide a superior signal to noise ratio in the microscope, whereas catalytic staining processes allow sensitivity to be increased by prolonging staining times. With most stains, selectivity is as important as sensitivity. Biological targets must often be stained differently from adjacent structures before identification is possible. Consequently an understanding of the mode of action of a staining method requires answers to two key questions:

   

  (i) Why does anything stain?

  (ii) Why doesn’t everything stain in the same way?

   

  This chapter provides answers to these questions. Although factors controlling sensitivity and selectivity are numerous, the same physicochemical phenomena are important whatever the staining methodology. Thus electric charges carried by dyes and biopolymers determine the selectivity of acid and basic dyes, and also influence the action of fluorescently labeled antibodies. The sizes of dye ions can control staining patterns, and the sizes of labeled antibody molecules critically influence sensitivity in the immunostaining of resin sections.

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  eBook - PDF

  Survey of Biological Progress

  Volume 2

  • George S. Avery, E. C. Auchter, G. W. Beadle, George S. Avery, E. C. Auchter, G. W. Beadle(Authors)
  • 2013(Publication Date)
  • Academic Press

    (Publisher)

  Histochemistry BY FLORENCE MOOG Department of Zoology, Washington University, St. Louis, Missouri I. INTRODUCTION Histochemistry (or cytochemistry 1 ) is in a broad sense a branch of physi- ology. Within the purview of histochemistry come not only the structures contained in cells and cell groups, and the chemical nature of these struc- tures, but also the activities that are carried on by cellular components, and the integrations of such activities. The proper aim of histochemistry is to contribute to our understanding of vital phenomena by specifying and de- fining their material bases. The science of cytochemistry was actually in existence in the middle of the nineteenth century, when a few workers were making attempts to de- termine the nature of intracellular elements. Actually the Golgi network was discovered in living spermatocytes teased out in a neutral medium. Ironically enough, it was the perfection of the microscope, the microtome, and the techniques of fixation and staining, beginning in the 1870's, that throttled the infant science of cytochemistry. Classical cytology does of course have a minimal chemical component. Particularly, the affinity of various cell parts for acid or basic dyes, or for certain metallic ions, has always been appreciated. The emphasis in cytology, nevertheless, was on the empirical development of effective staining methods, without regard to their chemical basis. The fascinating yet superficial discoveries of cytology inevitably led to a desire for a deeper understanding of the nature of cells and tissues. Progress in this direction followed soon on the publication, in 1936, of Lison's Histo- chemie Animale. Here, within a single slender volume, it was made clear that numerous satisfactory methods already existed for the demonstration of proteins, fats, carbohydrates, various metallic ions, and other substances in tissues.

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  eBook - ePub

  Histological Techniques

  An Introduction for Beginners in Toxicology

  • Robert Maynard, Noel Downes, Brenda Finney(Authors)
  • 2015(Publication Date)
  • Royal Society of Chemistry

    (Publisher)

  CHAPTER 7

  Standard Staining Techniques

  Staining of histological sections is both an art and a science: neither aspect should be ignored. In this chapter the technical aspects of staining are considered; the scientific principles underlying staining are considered in a later chapter. We begin by describing the rather simple apparatus needed for staining small batches of sections. In most laboratories staining is often done using staining machines and these, when properly adjusted and used, can produce excellent results. The beginner should always start by staining his or her own material by hand. A series of standard techniques have been set out step by step. Attention has been paid to the use of the staining microscope. This allows the depth of stain to be judged and, as necessary, adjusted. None of the techniques described here are difficult, though some are lengthy and fiddly to get right. Practice is, of course, the solution to most problems that the beginner is likely to encounter. The haematoxylin and eosin (H&E) method has been considered in some detail. This is the most widely used of all histological staining methods and will almost certainly be used by the reader. Familiarity with the H&E method sometimes breeds contempt and not enough attention is paid to producing first-class H&E stained sections. The beginner is encouraged to practise his or her H&E staining until the procedure become automatic and the results uniformly excellent.

  7.1 Introduction

  A justifiable sense of achievement or relief may be felt when you have a series of sections ready for staining. Staining is the part of section preparation that is the most fun and certainly the most interesting. In this chapter the apparatus needed for staining is described and then a limited number of standard methods are set out in some detail. The number of methods has been deliberately restricted: there are enough for most studies where the histopathological effects of chemicals are being investigated. Many more techniques may be found in the works listed in the Bibliography.

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