Use Of Purification System For Hla Sequencing-based Typing

The human leukocyte antigen (HLA) system is comprised of a group of genes essential to immune function

. This group of genes encodes cell-surface antigen-presenting proteins on the outside of a cell, which identify the cell as a self cell or non-self cell.

The immune system seeks out non-self cells for destruction. Matching HLA types reduce the risk of transplant rejection or an adverse immune response. Solid organ transplantation and allogeneic stem cell transplantation using DNA isolation represent a common treatment for end-stage organ failure and several hematological and non-hematological malignancies. Matching the patient with an unrelated donor for HLA molecules significantly decreases the probability of graft rejection, exposure to any graft diseases, and transplant-related mortality. HLA type screening methods are categorized as low, medium, and high resolution, where sequencing-based typing (SBT) provides the highest resolution and is considered the gold standard of HLA typing.

The major advantages of HLA sequencing-based typing are:

"Able to cover HLA complexity and polymorphism

"Capable of detecting new, undefined alleles

"Easily automated for high throughput and data consistency

Need for Streamlining HLA-SBT Workflow using Purification systems

With the increasing cost of BigDye genomic research centers were seeking solutions for hastening their research and reducing the operational costs of the HLA-SBT pipeline. Incorporated BigDye Terminator decreases flexibility, can limit research to particular clean up protocols, and increases unexpected nucleotide base calls, which can be interpreted as a heterozygous base call. Un-incorporated BigDye Terminator increases testing flexibility by expanding the clean up options and decreasing the possibility of unexpected nucleotide base calls. Achieving a BigDye ratio under 1:4 in an appropriate elution buffer would result in high quality reads, high signal to noise ratios between 20-2000, low background noise, and a good "G" signal. M.D. Anderson had three major goals for streamlining the HLA-SBT pipeline:

"Increase sensitivity

"Decrease BigDye dilution

"Achieve ease of use via automation

By using DNA purification system, all the objectives are met, as it is based on SPRI (Solid Phase Reversible Immobilization) paramagnetic bead-based technology and is a patented purification technology. It is a simple and highly efficient means of magnetic-based nucleic acid isolation used extensively in the Human Genome Project. The purification process consists of immobilizing nucleic acids onto paramagnetic micro particles under specific buffer conditions.

Flexibility is added to the system through modification of the binding buffer, which alters the type and size of immobilized nucleic acids. Sample contaminants are easily removed without the need for the centrifugation or filtration, thus creating an automation-friendly format.

Reference - https://www.beckmancoulter.com/wsrportal/bibliography?docname=HLA_AN.pdf

by: Scott eric

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