Immediate Spin Crossmatch

Principle:

Immediate-spin crossmatch ISXM is an abbreviated compatibility testing that detects most ABO incompatibility.  AABB Standards restricts its use to specific situations as indicated in the Policy section below.

Policy:

  1. Applicability:
    1. ISXM may only be used when ALL of the following conditions are met:
      1. Negative antibody screen
      1. No history of antibodies
      1. No ABO/D antigen typing discrepancies
      1. Less than two (2) ABO/D determinations are on-file.
    2. Use the computer/electronic crossmatch whenever its specific criteria are met.
  2. Testing:
    1. ISXM consists of reacting an RBC suspension with patient/serum or plasma, centrifuging, and immediately reading for agglutination or hemolysis.  THERE IS NO INCUBATION STEP, NO USE OF ANTIHUMAN GLOBULIN AHG REAGENT.
    2. If any hemolysis or agglutination is detected, the crossmatch is incompatible:
      1. Repeat the ISXM.
      2. Repeat the patient ABO/D AND unit ABO/D typings.
      3. Repeat the antibody screen.
      4. Perform full AHG crossmatch AND AHG/enzyme antibody panels.
      5. Refer the case to senior technical staff or a Transfusion Medicine consultant for further instructions.
        1. Case review will be documented in Medinfo HIIG computer system.
      6. DO NOT RELEASE THE RBCS UNTIL SO INSTRUCTED BY SENIOR STAFF.

References:

  1. Technical Manual, Current Edition, AABB, Bethesda, MD, USA
  2. Standards for Blood Banks and Transfusion Services, Current Edition, AABB, Bethesda, MD, USA
  3. Guidelines to the Preparation, Use, and Quality Assurance of Blood Components, European Committee (Partial Agreement) on Blood Transfusion (CD-P-TS), Current Edition

Antiglobulin AHG Phase Crossmatch

The antiglobulin phase crossmatch is the same indirect antiglobulin test IAT used for antibody screening, only instead of reagent RBCs we use the donor’s RBCs.  One question I like to ask my students is to describe the difference between the IAT and the antibody screen.  Most don’t realize they are the same thing, the same procedure, i.e. mixing plasma/serum with RBCs and then detecting a reaction using the AHG reagen

In Medinfo, the AHG phase crossmatch will be required unless the criteria for electronic crossmatch have been met (see my previous post on this topic).  In emergency mode, one can also do a class II release using immediate-spin to detect ABO incompatibility.  Medinfo will check the historical records for previous antibodies.

Notice that the process includes contingencies for room and even lower temperature conditions as well as immediate-spin.

I warn my students that we can make any tube, gel or glass-bead well react if we do not strictly adhere to the manufacturer’s recommendations.  The simplest example is to leave the reaction mixture to incubate longer than the time limit for LISS (e.g. 60 minutes).

If the antiglobulin phase crossmatch is positive (i.e. nonnegative reactions), then the unit is rejected or a least-incompatible crossmatch mode is activated.  The latter requires the transfusion medicine physician to specifically approve the allocation and release of this component.

The following is the Medinfo process I used:

To Be Continued

21/7/20

Antibody Titration including Software Processes

Processes and Software Building—Part 20

My practice across the globe has exposed me different rationales to performing antibody titration.  In my American training and practice (and also at international institutions following the American version of AABB accreditation), I only routinely performed titration of anti-D for Rh(D) hemolytic disease of the newborn and anti-A/anti-B for ABO-incompatible stem cell transplants AND ABO-incompatible renal transplants.

I have had heated arguments with some physicians who insisted they wanted titers for other antibodies.  The AABB Standards do not require this but leave it to the discretion of the Transfusion Service Medical Director.

In my entire career, I never worked in a blood bank or blood center which had optimal staffing or resources.  I focused on what was medically/technically necessary and even then still had shortages.  If performing a test will not change the clinical treatment, why perform it unless you are doing a research project!

Titration is a time-consuming, and until recently, a tedious manual task.  Recently some of the automated immunohematology analyzers offer a titration program.  We used the Ortho Vision Max which could perform both IgG and IgM titers within one hour—walk away!!  However, during that time, the titration procedure monopolized the analyzer.

The ABO-incompatible renal transplant program at HMC Qatar was modelled after Sweden’s Karolinska Institute.  However the latter site performed manual IgG and IgM titrations using Biorad/Diamed gels.

I did not have sufficient resources to commit staff to manual titration at HMC so I did a comparison study between the Ortho Max and the Biorad methods.  We were able to get good correlation and used the automated method for the transplant.

I am still biased against performing titrations for other antibodies.  I always ask, ‘Does the titration correlate with clinical severity?’  Unlike anti-D, antibodies such as anti-Kell and anti-c may be low titer but cause death.  Can anyone show me a definitive study that titers are useful except for transplants and Rh(D) hemolytic disease of the fetus/newborn?

Since the method was working well on the Ortho equipment, I next established an interface to Medinfo.  The test was performed separately for IgG and IgM antibodies.  Medinfo recorded the reactions in all the wells.  The last well showing a 1+ reaction was interpreted as the titer (e.g. if 1:64 were the last 1+ reaction, then the titer was 64 in Medinfo).

The Medinfo process is shown below.

20/7/20

Platelet and Plasma Allocation Rules

Processes and Software Building—Part 20

Medinfo software is rules-based so the institution may set its own custom rules for all processes.  One chooses a framework and then adds any additional rules it needs for optimization.  Turnkey systems do not offer this flexibility.

The rules for platelet and plasma components are much simpler than those for RBCs since usually we only consider ABO type.  There are two modes:  regular and emergency, the latter applying if not all the patient testing (including historical checking) is available.  The components, on the other hand, must meet all criteria before being considered for patient use.

Example rules for plasma follow:

For platelets, note that for adults and anyone else >= 20 kg, I gave any type of platelet pool or plateletpheresis component without regard to ABO matching.  With our production method, I did not give Rh immunoprophylaxis to females of child-bearing age receiving platelets from D-positive donors based on our clean (essentially RBC-free) Reveos automated production process.

Similarly, allocation rules for granulocytes, etc. could be made and enforced by the software.  Low-B-titer group A universal plasma would also be easy to implement.

To Be Continued:

19/7/20

RBC Antigen Matching and Software Processes

Processes and Software Building—Part 19

In the previous post I outlined how Medinfo handled antibody screening and identification.  This post reviews how antigen matching is used based on these results.

There are two modes, regular and emergency.  If the patient has not had at least two ABO/D determinations and/or does not have a recent antibody screen, then emergency mode must be selected with its own rules.  Otherwise, the regular mode applies.

Regular Mode:

In general, if there is a clinically significant antibody, an RBC unit which has not been matched for the corresponding antigen or has the corresponding antigen cannot be routinely selected.  However there is a hierarchy here also:

  1. Absolutely prohibited release—no one can override the logic (e.g. giving group O to a patient with anti-H)—not even the transfusion medicine physician can override this
  2. Restricted release—only certain staff can release the incompatible or untested unit (e.g. giving C-positive unit to someone with anti-C)
  3. Least-incompatible for WAIHA:  requires transfusion medicine physician approval
  4. Informational release:  authorized staff may release antigen-incompatible or untested unit but a pop-up menu appears and asks them to accept (e.g. Lewis untested unit in a patient with anti-Lea).
  5. Antigen-specificity matched—the usual mode for patients with antibodies

Examples of Regular Mode rules follow (these are not the complete lists but just provided to show the complexity of the process).

Emergency Mode:

This is much more restricted for selection of ABO/D and other antigen typings.  An example follows:

A similar hierarchy exits for platelet and plasma allocation and will be considered in a future post.

To Be Continued:

18/7/20

Antibody Identification and Software Processes

Processes and Software Building—Part 18

The blood bank is one of the last bastions of manual testing, especially for complex procedures.  Basic antigen typing, direct antiglobulin tests, and antibody screening adapt well for routine automation, but the actual antibody identification requires multiple methods and requires considerable experience to complete. At least today, a machine cannot replace the blood banker!!

One could take the approach to perform antibody panels in an automated system and record the results.  However, these are worthless unless you have the actual panel in hand.  Artificial intelligence to analyze the panel results is limited.  You still have to manually review everything.  In Medinfo, we could also scan the manually performed panels and add them to the record.

While I was at HMC, some of the hospital transfusion services elected to run panels automatically and then review the results manually.  We prepared a contingency to record the panel results but these by themselves could only be used with the actual red cell panels in use.

Examples of 11-R and 15-cell panel result importation follow:

What we absolutely had to do was enter the antibody specificity in a way that it could trigger our algorithms, i.e. our rules-based system of RBC allocation according the permissible blood types.  Each antibody specificity and each antigen typing had its own unique codeIt could not be entered free text!  Workflows follow:

Next, the antibody specificity test would trigger antigen typings for the correspond antigen and/or closely related antigens:

Based on the antibody specificity and antigen typings, a whole series of rules for routine and emergency-mode release were triggered.  This will be discussed in a subsequent post.

To Be Continued:

17/7/20

Extra Antigen Typings and Selection of RBC Units

Revision Date:  15/7/20 (replaces 9/6/20 post on Linked In and now recommends full extended Rh phenotype matching for sickle cell patients).

Principle:

Extended/extra antigen typing may mitigate alloimmunization in selected populations (chronically transfused such as sickle cell anemia, thalassemia, leukemias, lymphomas, etc.) and those patients already making RBC antibodies.  It can also help in discerning antibody specificities in patient workups where the antibody identification is unclear.  This policy is in addition to the current selection of antigen-negative RBCs when clinically significant antibodies are present (e.g. Kell-negative RBCs for patients with anti-Kell).

Policy:

  1. Perform extended Rh/Kell typing (CcEeK) in the following cases:
    1. Patient has a positive antibody screen during routine testing (unless typing has been previously performed and is showing in Medinfo Hematos IIG)
    2. All patients with hematopoietic neoplasia or related conditions: leukemias, lymphomas, myelodysplastic states, myeloproliferative states (polycythemia rubra vera, primary thrombocythemia, agnogenic myeloid metaplasia/myelofibrosis), aplastic anemia
    3. All patients with sickle cell anemia, thalassemia, sickle-thalassemia
  2. Selection of Donor Units in these situations:
    1. Match R1R1 when possible
    2. Match K-negative for K-negative patients
    3. Match rr for females <50 and children < 18 years
    4. There is no need to routinely match R1r, R2r, Ror, R1R2, or R2R2 genotypes with theses corresponding donor types except for sickle cell patients (item #5)
    5. Match exactly extended Rh phenotype and Kell for sickle cell patients (CcEeK): e.g. evenR2R2 for R2R2
  3. Hemolytic Disease of the Fetus/Newborn:
    1. If the neonatal eluate or mother at term shows a clinically significant antibody, phenotype the baby for the corresponding antigen (e.g anti-Fya  present, type baby for Fya)
  4. Extended antigen typing beyond Rh/Kell (e.g. k (cellano), Kpa, Kpb, Fya, Fyb, Jka, Jkb, MNSs, etc.) should be performed in the following cases:
    1. Sickle anemia, thalassemias, sickle-thalassemia
    2. Any antibody workup where the pattern is nonspecific
    3. Verify any antibody specificities by phenotyping the patient (e.g., if anti-M identified, do M typing)
    4. Cw phenotyping is not required unless anti-Cw is suspected.
  5. Genotyping may be considered if:
    1. Recently transfused patient and the extended typing is needed for antibody identification
    2. Ambiguous or unusual typing results (e.g. M-neg N-neg, lacking CcEe antigens by serologic typing)
  6. When NOT to phenotype:
    1. RBC transfusions within 3 months
    2. Previous phenotype in Medinfo Hematos IIG (unless the patient subsequently has had a stem cell transplant)
  7. Any other cases in which the Head, Transfusion Medicine or the other transfusion medicine physician requests additional typing.

Prepared By:

Zeyd Merenkov, MD, FCAP, FASCP

Senior Consultant/Division Head, Transfusion Medicine

Nursing Orientation from NGHA Riyadh

When I was affiliated with National Guard Health Affairs in Riyadh, my staff and I gave weekly new-arrival nursing orientations.  The attached PowerPoint file from 2004 shows the manual system in effect at the time, but it is still illustrative to acquaint new staff on the hospital blood bank as it relates to their nursing duties, including specimen collection, pick up and transfusion of blood components, and adverse effects of transfusion.

I wish to give full credit to Mr. Abdullah Al Khashan, who prepared this file in conjunction with Ms. Editha Durante, the Transfusion Clinical Resource Nurse.  The three of us used to rotate giving this lecture.  This is my version of their presentation (with some minor formatting changes).

Overwashing During Elution

I cannot emphasize enough proper technique in doing the washing during the elution process.  We are usually concerned about too little washing and thus possibly residual reactions in the last wash.  However, aggressive overwashing may remove the bound antibody resulting in a negative result.

Here is an example of anti-PP1Pk (alias anti-Tja).  The mother’s panel shows an antibody to a high prevalence/incidence antigen with negative autocontrol and no lability at enzyme phase:

The neonate’s DAT was weak positive at polyspecific and IgG monospecific phases.  An eluate was performed.  Here is the result after washing four (4) times:

Since 2 cells in the last wash were very weakly positive, the washing was continued for a total of 9 times with the following results:

Even then there was very weak positivity in one cell, but the eluate was negative.  We had washed away the attached antibodies.

Elution Indications and Software Implementation

Processes and Software Building—Part 17

The direct antiglobulin test DAT may be the first or last place that a delayed hemolytic transfusion reaction can be detected.  In my practice, I always performed it the first time I encountered a patient with a positive DAT.  If the subsequent DAT testing is of the same strength and there is no change in the clinical status of the patient, then I might empirically repeat the elution after 14 days, i.e. just when new antibody emergence may be detected.

In summary, my criteria for elution after a positive DAT are:

  1. First patient encounter with a positive DAT
  2. At least 14 days since the previous elution
  3. DAT strength has increased since the previous exam (at least 1+ increase in strength in either IgG or C3)
  4. Patient shows evidence of hemolysis
  5. Suspected cases of drug-related hemolysis
  6. Transfusion Medicine physician specifically orders it otherwise

In the software processes, the following parameters were recorded:

  1. Technique of elution (acid glycine, dichloromethane, digitonin, ether, etc.)
  2. Number of washes (too many may remove the antibody from the RBCs)
  3. Last wash result (must be negative to accept conclusion)
  4. Manufacturer of the elution kit (if any)
  5. Elution result based on panel testing

Note:

  1. All indeterminate or positive eluates would trigger an antibody identification.
  2. All results would be reviewed by me or the covering Transfusion Medicine physician.  A comment would be entered into the results that could be viewed by the clinician either directly from Medinfo or through the hospital information system.

If the DAT was only positive for complement, it was up to the Transfusion Medicine Consultant to decide whether to proceed with elution.  Personally, I would usually proceed because the eluate is more concentrated and may occasionally detect an antibody that was not noted in the patient’s plasma.

The attached Medinfo workflow shows the process designed by me for use at HMC Doha.