Clinical Immunogenetics Laboratory (2024)

Description of HLA

HLA stands for “Human Leukocyte Antigen.” These antigens are protein molecules we inherit from our parents. Together, these molecules make up your HLA type. We currently know about more than 2500 different HLA molecules. It is very important to determine your HLA type before you have a stem cell transplant, and we can do this by taking a sample of your blood or other body tissue cells.

HLA testing will also determine the HLA type of anyone who may donate stem cells to you. It is important in stem cell transplants to see how closely the HLA type of the transplant patient matches the HLA type of the stem cell donor. The HLA “match” is the number of HLA molecules that any two people have in common for stem cell transplantation. HLA matching is usually based on 10 HLA molecules. The more molecules two people share, the better the match. When two individuals share the same HLA molecules, they are said to be a good “match.” That is, their immune systems will not see each other as “foreign” and are less likely to attack each other.

The most likely place to find an HLA match between two people is among siblings (that is, brothers and sisters who have the same mother and same father). If two siblings inherit the very same HLA molecules from both parents, they are said to be an “HLA identical match.”

You have a 25 percent (1 in 4) chance of being an HLA identical match with your sibling. Why? Because there is a basic rule in HLA inheritance: you have a 25 percent chance of inheriting the same HLA molecules as your sibling, a 25 percent chance of inheriting none of the same HLA molecules as your sibling, and a 50 percent chance of inheriting half of the same HLA molecules as your sibling.

However, two unrelated people can just happen to be a good HLA match, too. Although it is less likely, it is possible that you could have some of the same HLA molecules as someone you don’t even know.

Request HLA Typing

We typically receive HLA typing requests from treating physicians, for example, your primary oncologist. While we can certainly discuss HLA typing with you directly, we recommend that your physician be involved to work with us.

Upon receiving your request, our Intake department will work with your local physician to determine three things:

1. What is the clinical urgency? This is based on your clinical condition.
2. What is your treatment plan?
3. Is searching for an alternate donor being considered if there is no family match?

After determining the items above:

• We contact you to explain the HLA typing process and collect insurance and family member information.
• We contact your insurance carrier for financial clearance and communicate financial issues to you to ensure you are aware of any financial implications from HLA typing.
• If we need to discuss altering the HLA typing strategy based on any of the information collected, our Clinical Coordinator Physician will contact your local physician and/or you.
• Our Clinical Immunogenetics Laboratory (CIL) will send instructions on how to collect and send specimens for HLA typing to your local physician.

If you want to discuss HLA Typing, feel free to contact our Intake department. The Intake department hours of operation are Monday through Friday from 8:30 am to 5:00 pm (PST) and our main phone number is 800.804.8824 or 206.606.1024.

Chimerism Testing

The Clinical Immunogenetics Laboratory offers chimerism testing of patients after HSCT (and for certain specific protocols before transplant) to determine recipient/donor origin of nucleated hematopoietic cells.

Chimerism Testing/Engraftment Analysis

A Chimera was a creature in Greek mythology usually represented as a composite of a lion, goat, and serpent. Contemporary use of the term “chimerism” in hematopoietic cell transplant derives from this idea of a “mixed” entity, referring to someone who has received a transplant of genetically different tissue. A test for chimerism after a hematopoietic stem cell transplant involves identifying the genetic profiles of the recipient and of the donor and then evaluating the extent of mixture in the recipient’s blood, bone marrow, or other tissue.

Chimerism testing (engraftment analysis) by DNA employs methodology commonly used in human identity testing and is accomplished by the analysis of genomic polymorphisms called short tandem repeat (STR) loci. These loci consist of a core DNA sequence that is repeated a variable number of times within a discrete genetic locus. The term STR, also referred to as microsatellites, relates to the number of base pairs of a tandemly repeated core DNA sequence which ranges from 2-8 base pairs in length. These loci exhibit alleles that may differ in length between individuals and are inherited as codominant Mendelian traits. STR loci have been identified throughout the human genome and some loci have more than 25 alleles.

DNA sequence information within the conserved flanking regions of the loci is used to create oligonucleotide primer pairs for the STRs. These primers are used in PCR (polymerase chain reaction) amplification of test samples. This technique can amplify the STR sequence as many as a billion times, providing material that can be separated with an electrophoretic gel or by capillary electrophoresis (CE). Genotyping is done by evaluation of the DNA fragment sizes. Reference to an allelic ladder may be used for exact identification of STR alleles.

The PCR-based STR/CE system has several advantages over other methods of analysis. The amplification of multiple STR loci can be combined (multiplexed) in a single tube, permitting analysis of up to sixteen loci in one reaction. Since minute amounts of DNA are required, samples with low cell numbers can be used, and the small size of the STR alleles even makes it possible to use degraded DNA samples. The digital data facilitates analysis and archiving, and the CE process is both fast and cost effective. PCR amplification and analysis of STR loci provides a rapid and reliable method for the evaluation of engraftment status in the stem cell transplantation setting.

Currently used technology allows the co-amplification and three-color detection of sixteen loci which are subdivided into 3 sets of 5 or 6 loci that exhibit amplified fragments with non-overlapping size ranges.

During the PCR amplification step, the amplified fragments are labeled with fluorescent dyes. After PCR amplification, the samples are processed on a capillary electrophoresis (CE) system.

Data analysis is facilitated by a fragment analysis software which sizes the DNA fragments using an internal lane standard run with the sample and assigns genotypes by comparison to an STR allele ladder included in the CE run. This provides distinct STR genotypic profiles for the donor and for the transplant recipient. STR loci that are polymorphic (i.e., informative) between these individuals are used to assess relative amounts of recipient and donor DNA in the post-transplant sample.

Samples tested may be from any material containing DNA, including bone marrow, peripheral blood, solid tumors, epidermal tissue, hair follicles, buccal swabs, and fractionated cell subsets. Since PCR amplification of a sample is routinely performed with less than 2 ng of genomic DNA (equivalent to approximately 300 cells), chimerism testing by this method can be successfully performed even for patients with graft failure, severe leukopenia, or from hematopoietic cell subset fractions. STR analysis has been used to evaluate the engraftment status of patients who have received a hematopoietic cell transplant including patients receiving double cord blood donor units or a second transplant from a different donor; as well as to confirm the genetic identity of putative identical twins, and to detect in-utero derived maternal cell engraftment among patients with a diagnosis of Severe Combined Immunodeficiency (SCID). STR/CE analysis is a rapid, reliable, accurate and reproducible procedure.

Clinical Indications for Chimerism Testing in Hematopoietic Cell Transplant

Routine post-transplant documentation of the donor/recipient origin of white blood cells in peripheral blood and/or marrow. Documentation of engraftment may include testing lineage-specific cell subsets, such as CD3 positive T-cells and CD33 positive myeloid cells.

• Evaluate donor/recipient cells in patients with inadequate marrow function.
• Define whether recurrent or new malignancy has originated from recipient or donor cells.
• Assess prognostic risks of rejection and recurrent malignancy.
• Document the persistence of donor cells post-transplant in patients with recurrent disease or prior to donor lymphocyte infusion (DLI).
• Evaluate whether graft rejection has occurred in recipients that are candidates for a second transplant.
• Differentiate the origin of donor cells in recipients who have received a second transplant with a different donor or a transplant with double cord blood units.
• Detect the presence of maternal derived cells in patients diagnosed with Severe Combined Immuno-Deficiency (SCID).
• Verify genetic identity of putative identical twins.

Literature Cited/Recommended Reading

Bryant E, Martin PJ. Documentation of engraftment and characterization of chimerism following hematopoietic cell transplantation. In: Forman S, Blune K, Thomas ED, eds. Hematopoietic cell transplantation. 2nd ed. Malden, MA: Blackwell Science, 1998.

Bultler J. Forensic DNA typing. Elsevier Academic Press.