cRF

The Calculated Reaction Frequency (CRF) was introduced to overcome problems with the previous Panel Reactivity (PRA) system which relied on the composition of the testing panel to give the percentage reactivity. This was however flawed as the panel composition changes between tests and may have an antigen frequency that does not reflect the donor population. Developed first in the USA and adopted by UNOS in 2006 and later in the UK, the CRF is based on a calculation of the reaction frequency of a listed set of unacceptable mismatches for a patient, against a panel of 10,000 recently added deceased donors. The CRF therefore gives a measure of the chances of a patient finding a compatible donor in the donor pool.

The use of CRF based on a set of listed unacceptable mismatches for each patient is made much easier by the wide adoption of solid phase assays such as Luminex. Luminex assays, especially those involving the use of single antigen beads (SABs) allow fine specificity definition and allows the strength of the reactions (MFI) to be used to assess immunological risk and help decide whether or not a specificity should be listed.

In the UK, patients with a CRF of 100% are prioritised into Tier A for kidney and pancreas allocation in the newly updated organ matching scheme. All other patients in Tier B are allocated points for high matchability scores which helps improve their chances of getting a offer.

Matchability

Matchability is a measure of how easy or difficult a patient is to match for an organ. In the UK it is calculated based on blood group, HLA type and HLA antibodies. It gives an estimate of the number of ABO identical, HLA compatible donors within a 10,000 donor pool a given recipient is matched with.

Matchability is scored on a scale from 1 -10, where 1-3 represent easy to match, 4-6 are moderate and 7 -9 are difficult. A matchability score of 10 denotes very difficult to match. In the UK, patients with a matchability score of 10 are prioritised into Tier A when deceased donor pancreas and kidneys become available.

HLA Matchmaker

It has long been known that antibodies do not bind to the whole of the HLA antigen but rather to specific epitopes on the antigens. Each HLA antigen potentially has many sites or epitopes that can bind antibody. These epitopes may be private to a given HLA antigen or they may be shared by more than one HLA antigen, i.e. they may be public. In recent years it has become possible to identify the polymorphic amino acid residues that define many of these public and private HLA epitopes. This has enabled the development of laboratory based and in-silico strategies for assessing HLA compatibility at the epitope level.

One laboratory based strategy for defining epitopes has been described by El-Awar and Terasaki et al. They have used single antigen beads (SAB’s) to define a set of 103 epitopes on HLA class I antigens. Their assays were performed with mouse monoclonal antibodies directed against HLA as well as with anti-HLA from sensitized patients and multiparous women. Where alloantisera was used, is was first absorbed out using SAB’s or cell lines before being eluted for testing in a Luminex assay.

An example of an in-silico assay is that proposed by Kosmoliaptsiset et. al. They have described an epitope definition system based on interlocus and intralocus comparison of patients and donors to identify amino acid differences but also crucially, they have carried out an assessment of the physiochemical properties of the amino acid mismatches and the role these may play in clinical outcome. They have applied this scheme to kidney transplantation in the UK.

The most widely known in-silico strategy in the field of HLA is however the HLA Matchmaker algorithm (http://www.hlamatchmaker.net) as proposed by Duquesnoy. Two versions of HLAMatchmaker have been described, the ‘triplet’ and ‘eplet’ versions. In the triplet version, epitopes are defined by linear sequences of triplets of amino acid residues in alloantibody-accessible positions of the HLA molecules (the a-helices and b-loops of the protein chain). The eplet based version of HLAMatchmaker defines epitopes as those residues located within a diameter of about 3-3.5 Å around a non self residue. Although some of these eplets consist of amino acids in linear sequences as described in the triplet model, many consist of amino acids from discontinues regions of the sequence, brought together by the folding of the molecule. In HLAMatchmaker, Donor-Recipient HLA compatibility is based same intralocus and interlocus comparisons of triplets or eplets.