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How do reagents contribute to IHC consistency?

Automated instruments aim to achieve consistency through controlled procedures and a reagent menu that offers quality-controlled manufacturing to minimize batch variation.

Concentrates or RTU antibodies?

When selecting antibodies, there are two main options to consider; a concentrated format or a pre-diluted, Ready-To-Use (RTU) format.

Concentrates are flexible, have a lower initial purchase price, and can generally be used within any staining system, both automated and manual, subject to the manufacturer’s recommendations. The working dilution of concentrates can be optimized to balance cost, staining time and quality. Given the large working dilution range, concentrates can be varied at any time to accommodate changes to laboratory practice or for multiple protocols for a particular antibody. However, concentrates do require preparation time and validation. As there is no definitive way of determining the properties and stability of a diluted antibody without well controlled and executed studies, staining quality may be compromised as subtle deteriorations may not be noticed.

The advantages of RTUs include increased laboratory efficiency, better quality control and easier reagent management. They eliminate time spent on working dilution, preparation and the time to validate the assay. Consistency is enhanced with run-to-run variation reduced, especially in conjunction with automated stainers and associated detection systems. With a defined number of tests and manufacturer-verified expiry, RTUs simplify antibody management. Additionally, RTUs can contribute to laboratory growth by making it easy to adopt new antibody assays as the amount of validation work is greatly reduced.

What makes a good detection system?

Detection systems for automated instruments provide quality controlled reagents associated with a verified protocol that achieves quality, consistent staining in the shortest possible time. These detection systems often include most reagents required for detection of multiple targets, for example antibodies or probes. The two most common detection systems used are the avidin‑biotin method or polymer-based detection.

The avidin-biotin method is well known and understood and is generally considered adequate. The newer polymer detection systems have the advantage of being biotin free and having far greater sensitivity. Polymer systems vary between manufacturers but the basic concept involves antibodies and enzymes conjugated to a polymer chain allowing multiple binding sites for increased sensitivity. Modern automated staining systems take advantage of the benefits of polymer detection to produce very fast, highly sensitive IHC staining. This helps laboratories reduce both staining costs and turnaround times.

Polymer detection systems allow multiplexing with multiple colored chromogens. This provides a high contrast stain visualizing more than one target on a section.

A vision for high-quality IHC and ISH.

At Leica Biosystems our vison is to advance cancer diagnostics and improve lives. One way we can achieve this vision is by helping improve staining quality. As we recognize that IHC and ISH quality doesn‘t begin at the stainer, this series looks at many different aspects of staining quality, and considers how future tests will influence improved diagnosis.