Stainers / Stains

Troubleshooting Histology: Double Barrel Style – H&E and Special Stains

Why even bother to learn about troubleshooting? Wouldn’t it be more interesting to listen to one of the presentations on molecular technology? Possibly. But what would you do if your pathologist brought you a slide from the first batch of today’s H&E’s that looked sub-optimal to review alongside an optimal slide stained the previous day? …

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Validating Your New Stainer According to CAP and CLIA Requirements

Congratulations on purchasing a new autostainer! There are three parts to the start-up process:  Instrument Verification, Stain Protocol Optimization, and Validation of the Staining Protocols.  Instrument Verification – see CAP All Common Checklist (06/04/2020) COM.40350 Optimally, the company you purchase the instrument from should provide an operator manual; provide verbal procedural instructions; and test the …

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#7 IHC Double Staining

The DNA of a nucleus directs the actions of the cell by instructing it to manufacture specific proteins.  This “instruction” is sent from the nucleus into the cytoplasm by “messenger RNA” (mRNA).  The mRNA exits the nucleus to bring the “message’” into the cytoplasm, binding with ribosomes to manufacture proteins.  Figure 1 illustrates this principle. …

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#6 IHC Nuts and Bolts

Procedures

Immunohistochemistry Educational Series In previous installments of the IHC Educational Series, we discussed the specifics of the basic procedures found in the IHC laboratory: immunofluorescence, immunohistochemistry and in situ hybridization. Even though these are different procedures, they all require the following support procedures.Whenever a new antibody (or probe) is ordered for use by a pathologist …

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#5: In Situ Hybridization

In situ hybridization

In the previous segments of the IHC Educational Series, immunofluorescence and immunoperoxidase methods were discussed.  In both of these methods, antibodies are used to bind to and localize protein antigens in tissue sections.  Additionally, the differences between the use of polyclonal and monoclonal antibodies had been described. Another immunohistochemical method is “in situ hybridization”. The …

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#4: Antibodies

In the previous units of this series, the immunohistochemical (IHC) methods of immunofluorescence and immunoperoxidase were described and discussed.  These methods use antibodies to localize cellular proteins in tissue sections, which can then be visualized in either a dark field fluorescence microscope or routine bright field microscope.  It is important to understand antibody structure and …

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#3: Immunoperoxidase

In the previous segment, immunofluorescence methods were discussed.  These methods use antibodies labelled with fluorescein to directly localize cellular proteins in tissue sections, which can then be visualized in a dark field fluorescence microscope.  Despite the advantages of this method, science is always in search of newer methods to gain more knowledge.  Thus in 1970 …

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#2 Immunofluorescence

Before we discuss immunofluorescence, we need to know what fluorescence is.  There are certain substances, composed of molecules that will emit light when irradiated by a short wavelength, such as X-rays or ultraviolet (UV) light.  The emitted light is of a longer wavelength which has a lower energy.  Two substances that are used in microscopy …

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#1 Introduction to Immunohistochemistry

Introduction to Immunohistochemistry

Anthony van Leeuwenhoek is credited with being the first person to use a microscope to view tiny “animacules” in 1674. At approximately the same time, Robert Hooke used a similar microscope to view thin slices of cork. The structures that he observed resembled the tiny cells that monks lived in at the local monastery, so …

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The H&E Stain: Far From Routine Part 2

This blog is a followup to the previous article “The H&E Stain: Far From Routine”.  In that article, the basics of the routine hematoxylin and eosin (H&E) stain were discussed.  Now we shall discuss how to trouble shoot the routine H&E and how to ensure a high quality stain, once you have worked with your …

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