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The human leukocyte antigen (HLA) system plays an important role in the regulation of the immune system; HLA recognizes cells as "non-self" or "self" to prevent the body from attacking its own cells.

For transplant rejection, when the donor cells are not recognized as "self" by the recipient, the HLA system plays a key role. It is known to cause some autoimmune diseases, including rheumatoid arthritis and multiple sclerosis.

HLA typing is a genetic test commonly used in diagnostic laboratories around the world. It uses molecular or serological methods to find recipients and donors with closely matched HLA patterns, thereby minimizing the risk of transplant rejection.

If the donor's HLA is different from the recipient's HLA and triggers an immune response against the donor, it will lead to cell death.

This article will discuss the many different HLA typing methods in use and how the ASSIST PLUS pipetting robot automates multi-channel pipetting to free up time for laboratory personnel and improve the reproducibility and accuracy of results.

Antibodies are used to distinguish different variants of HLA antigens in HLA serotyping. Donor serum is mixed with a set of HLA-specific antibodies to determine which HLA patterns the cells express and test for immune response.

Serotyping using final cross-matching allows screening of pre-existing antibodies to compare the HLA of the recipient and donor before transplantation. During this process, the patient's serum is mixed with the donor's lymphocytes.

Both applications use a special plate called Terasaki tray. The tray contains 72 or 60 holes, each with a capacity of approximately 10 µl; mineral oil is used to prevent evaporation of the sample dispensed on the tray.

Even the most experienced laboratory technician will find it tricky to accurately move these small volumes into the oil in the center of each well. Despite the unique dimensions of the Terasaki tray, the use of ASSIST PLUS and VOYAGER adjustable tip spacing pipettes can automate the process.

Every time, automation is used to ensure the accurate positioning of the pipette tip, so that small volumes can be accurately and precisely dispensed at a consistent speed and immersion depth.

INTEGRA provides a specially designed adapter for this application to maximize efficiency, allowing users to increase throughput and unattended time by continuously processing up to two Terasaki boards.

The reactivity between the donor HLA and the recipient antibody can also be determined by serotyping as an ELISA. Luminex technology is a bead-based immunoassay system. Compared with standard ELISA, it can detect up to 100 analytes at the same time; this provides higher sensitivity and specificity for HLA testing.

Therefore, it has become the clinical standard for identifying donor-specific HLA-specific antibodies in solid organ transplantation worldwide.

Luminex and ELISA tests can be efficiently processed with VIAFLO electronic multichannel pipettes because they are based on multi-well microplate tests, which are usually performed in 96-well plates.

By clicking the pipette into ASSIST PLUS, you can free up valuable time in busy clinical laboratories and minimize differences in results between operators, thereby automating the ELISA step.

The ASSIST PLUS pipetting robot can easily and automatically complete all the tedious pipetting steps in the immunoassay, from stopping the reaction, adding the sample, and transferring the sample to the coated plate.

When it became clear that there are HLA variants that cannot be serologically distinguished but can be viewed by looking at underlying genes, a clinical molecular typing strategy was developed.

Next-generation sequencing (NGS), PCR sequence-based typing (PCR-SBT), PCR sequence-specific oligonucleotide probes (PCR-SSOP), and PCR sequence-specific primers (PCR-SSP) are some sequence-based Protocol in use for HLA typing.

All molecular HLA typing methods, including those listed above, first isolate the nucleic acid from the sample, and then they use PCR to amplify the target region of interest-certain HLA exons, full-length genes, or complete HLA regions.

It takes multiple PCR reactions to type a sample because the HLA system is a large gene complex spanning 3,600 kilobases.

INTEGRA provides the best system for sample nucleic acid extraction using the ASSIST PLUS pipetting robot. The system can also automatically perform PCR settings for molecular HLA typing, thereby reducing the overall cost of each sample and significantly reducing manual operation time.

Even PCR settings in 384-well plates can be easily automated, enabling efficient typing of large numbers of samples. By using ASSIST PLUS to automate the VOYAGER adjustable tip spacing pipette, multiple parallel channels can be used to add samples stored in test tubes to the plate, ensuring precise and accurate handling of small volumes.

The SureFlo™ anti-sealing array of the reservoir provides the lowest dead volume available on the market, which means that combining this setup with INTEGRA's multi-channel reagent reservoir can minimize the consumption of expensive reagents.

The user can ensure that the sample stays cool during the preparation of the assay by using the adaptor for the cooling of the reservoir and 96-well or 384-well PCR plate.

For SBT and NGS methods, the amplicon must be quantified and purified after the PCR step-this is usually accomplished by magnetic bead purification. Users can use ASSIST PLUS's flexible platform settings to install magnetic plates during the purification process to automatically perform tedious pipetting steps.

Optimal pipetting settings can maximize sample recovery, including tip angle, pipetting speed, and tip immersion depth. Ensure the consistent positioning of the needle tip in the sample hole, minimize the risk of interfering with beads or bead residue, and provide high sample yield and purity for downstream sequence analysis.

If the donor and patient do not match, inaccurate liquid handling or human error in the HLA typing process may lead to serious consequences. These risks can be minimized by automating the workflow.

Compared with manual workflow, using ASSIST PLUS pipetting robot means pipetting small volumes with higher precision and accuracy, which is especially important when samples may be precious and small.

By combining ASSIST PLUS with one of INTEGRA’s 25 electronic multi-channel pipettes, users can easily switch between different processes and applications according to their workflow needs, not just one HLA typing method , The pipette provides a variety of volumes and pipetting channels.

It is even possible to automate the processing of Terasaki trays or tube racks through the VOYAGER adjustable tip spacing pipette on ASSIST PLUS, thereby improving work efficiency. Diagnostic laboratories and blood transfusion and transplant centers wishing to enhance HLA typing capabilities will find ASSIST PLUS to be a multifunctional tool.

INTEGRA provides innovative solutions for liquid handling and media preparation applications to meet the needs of customers in research, diagnostics and quality control laboratories.

Their instruments and plastic consumables are developed and manufactured in Zizers, Switzerland and Hudson, New Hampshire, USA. In order to maintain close contact with customers, they have direct sales and support organizations in North America, the United Kingdom, France and Germany, and have a network of more than 100 well-trained distribution partners worldwide.

In recent years, they have focused on developing new and technologically advanced series of handheld electronic pipettes that are easy to use and meet the ergonomic needs of customers. Today, they proudly offer the widest range of electric pipettes on the market, ranging from single-channel pipettes to 384-channel benchtop instruments.

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Last update: November 10, 2021 at 10:07 am

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INTEGRA Biological Sciences. (2021, November 10). Automated human leukocyte antigen (HLA) workflow. News-Medical. Retrieved from https://www.news-medical.net/whitepaper/20211101/Automating-the-human-leukocyte-antigen-(HLA)-workflow.aspx on November 25, 2021.

INTEGRA Biological Sciences. "Automated Human Leukocyte Antigen (HLA) Workflow". News-Medical. November 25, 2021. <https://www.news-medical.net/whitepaper/20211101/Automating-the-human-leukocyte-antigen-(HLA)-workflow.aspx>.

INTEGRA Biological Sciences. "Automated Human Leukocyte Antigen (HLA) Workflow". News-Medical. https://www.news-medical.net/whitepaper/20211101/Automating-the-human-leukocyte-antigen-(HLA)-workflow.aspx. (Accessed November 25, 2021).

INTEGRA Biological Sciences. 2021. Automated human leukocyte antigen (HLA) workflow. News-Medical, accessed on November 25, 2021, https://www.news-medical.net/whitepaper/20211101/Automating-the-human-leukocyte-antigen-(HLA)-workflow.aspx.

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