Immunohematology - Laboratory F1: Compatibility Testing PDF

[MLS 416-LEC] Immunohematology - Laboratory F1: COMPATIBILITY TESTING Professor: Ichiro Ordaneza, MD Date: April 24, 2024 COMPATIBILITY TESTING Also called Pretransfusion testing ○ Processes done before the actual transfusion process Series of serologic and non-serologic protocols/ Testing procedure...

[MLS 416-LEC] Immunohematology - Laboratory F1: COMPATIBILITY TESTING Professor: Ichiro Ordaneza, MD Date: April 24, 2024 COMPATIBILITY TESTING Also called Pretransfusion testing ○ Processes done before the actual transfusion process Series of serologic and non-serologic protocols/ Testing procedures with the ultimate objective of preventing an immune mediated hemolytic transfusion reaction 3 Categories PURPOSE: To select blood components that will not cause harm to the recipient and will have acceptable survival when transfused Main goal of MedTechs: Ensure safe transfusion of blood units between the Donor and the Recipient If properly performed, compatibility tests will confirm ABO compatibility between the component and the recipient and will detect the most clinically significant unexpected antibodies. Take note that i could not guarantee normal survival of transfused red blood cells in the circulation of the patient There are still potential risk involved in the procedure if this type of therapy is to be considered ➔ ➔ ➔ ➔ ➔ Components of Compatibility Testing Proper specimen collection Reviewing patient transfusion history ABO, Rh and antibody testing (Screen/ID) Crossmatching Actual transfusion It is our responsibility and not the patient, to ensure correct and uneventful transfusion process even right from the patient identification Pretransfusion testing BEGINS and ENDS with the proper IDENTIFICATION and COLLECTION of the patient’s samples Preanalytical procedures Serological testing Postanalytical procedures PRE-ANALYTICAL PHASES Patient identification Specimen collection Review of patient history Patient Identification Major cause of transfusion related fatality (Clerical error) – resulting in incorrect ABO groupings and transfusion of incompatible blood ○ Greatest threat to safe transfusion therapy ○ Misidentification of patient Must confirm recipient’s ID from bracelet ON the patient – Held ID is not allowed, it must be attached to his/her body ○ Full patient name (first and last name) ○ Date of birth ○ Hospital number ○ Name of physician For OUTPATIENTS it is important to ask for the REQUEST FORM that must coincide with the LABORATORY REQUEST and asking him/her to state his/her FULL NAME and DATE OF BIRTH (For confirmation/coherency) ○ NEVER ask the WATCHER, ALWAYS ask the PATIENT If incoherent ask the nurse Sample Identification Patient name (Full) Hospital number Physician Date and Time of collection Phlebotomist Initials All of this should be on the request form and the sample Copy for: HAW, SPENCER O. | 1 Phlebotomist’s initial is important because so that when anything happens we can keep track of it The REQUEST FORM and the LABEL ON THE SAMPLE should coincide with each other Make sure that your handwriting is legible Specimen Tubes Pink Top - EDTA For plasma, ANTICOAGULANTS may inactivate the complement system, this is due to the calcium chelation activity of EDTA, so that some antibodies may not be detected. But plasma (anticoagulated blood) is more used today due to ease of handling because the benefits outweigh the potential issues. Red Top - No additives Clotted Red Blood Cells may require additional WASHING STEPS to minimize interference in test interpretation For serum, its DISADV. Is that there could be formation of small fibrin clots which may be difficult to distinguish from TRUE AGGLUTINATION Take note that upto 10 mL of blood may be collected for all serological test inside the blood bank Specimen Collection Blood Collection and Handling Tubes: Use lavender or red top tubes (EDTA or no additives) Hemolysis: Avoid hemolysis during venipuncture (can affect results). ○ True hemolysis – due to complement activation ○ Sample hemolysis – due to improper blood collection Labeling: Label samples at bedside after collection (not pre-labeled). ○ After blood collection, that is the time that you must do your labeling ○ Should be inside the patient’s room or the patient is still present Documentation: specimens should be documented in order to “backtrack” in the case of an error Avoiding Errors Hemolyzed Samples: Can mask true antigen-antibody reactions. IV Line Contamination: Unacceptable, dilutes sample and gives erroneous results. ○ Stop IV line 5-10 minutes before draw in non-ICU patients (discard first 10ml). ○ Collect blood below IV line in ICU patients (consult nurse). ○ If contamination is suspected, redraw the sample. Sample Processing and Storage Serum: Separate serum from RBCs ASAP after clotting. Testing: Perform tests within 72 hours for accurate complement detection. Storage: Store samples at 1-6°C for up to 72 hours for future testing. Transfusion Reaction Sample Retention: Keep patient and donor samples for 7 days post-transfusion for error investigation. If a transfusion reaction occurs… The patient sample and the segment from the donor unit must be retained post-transfusion for at least 7 days – because if such errors arise, we now have a sample that we can use for investigating the error. Getting The History Current ABO/Rh type Significant antibody detected Significant adverse events Look at recipient’s records for any prior unexpected antibodies Previous transfusion reactions Special transfusion requirements Many antibody titers decrease to undetectable levels; Between 30-35% of antibodies reach undetectable levels within one year and nearly 50% become undetectable after 10 years. One example is the KIDD BGS, which is notorious for causing a delayed type of hemolytic transfusion reaction Resolve any issues in patient history before proceeding to the next steps Copy for: HAW, SPENCER O. | 2 SEROLOGICAL TESTING It is in this phase where the samples we collected will be tested. ABO/Rh type Antibody screening/ detection/Identification Crossmatch You cannot proceed immediately to antibody detection or Cross Matching without first performing ABO/Rh typing (1) ABO/ Rh Typing The MOST CRITICAL pretransfusion serologic test; because this blood group continues to be the most important BGS in transfusion medicine due to their immunogenicity. In the ABO typing, the forward and reverse MUST match ○ Resolve any discrepancies if there are any ○ Can be performed through: Slide, Tubes, Gel Technology, Solid-Phase RBC adherence column ○ If the ABO discrepancy cannot be resolved immediately and immediate transfusion is required – you must give Group O negative RBCs (First line blood transfused in a emergency setting) In the Rh typing, the control must be NEGATIVE ○ To detect the presence/absence of D antigen ○ A control must be present in detecting for weak D antigen using IAT ○ If test control are positive the test is INVALID ○ Rh negative recipients should receive Rh negative RBC to prevent production of anti-D alloantibodies Remember that if a patient has a weak-D antigen, the patient is still labeled as Rh Positive; this poses no risk for developing anti-D because the antigen sites are only reduced, they still possess D antigen. Mosaic D individuals however should be labeled as Rh negative as recipient and Rh positive as donor because of the missing D epitopes. Both of these will indicate what type of blood should be given ○ Immediate Spin (IS) ○ 37 deg C (LISS) ○ Antihuman Globulin (AHG) Regardless of the phase, transfusion of antigen negative red blood cells is the transfusion requirement if the patient has antibodies against that antigen. ○ If an antibody is present, units negative for the antigen must be given Clinically significant antibodies are reactive @ 37 deg C ○ But some antibodies that are of the IgM type may be detected at lower temperature (@RT) Proceed to the crossmatch (3) Crossmatching Major Crossmatching Minor Crossmatching PS-DR PR-DS (Px Serum - Donor Red cells) Routinely performed in labs (Px Red cells - Donor Serum) Not required by AABB since 1976 DO NOT INTERCHANGE! Purpose: Prevent transfusion reactions Increase in vivo survival of red cells Double checks for ABO errors Another method of detecting antibodies A non-reactive crossmatch indicates the donor units are compatible and safe for transfusion (THIS IS WHAT WE WANT!) (2) Antibody screen and/ or Identification The antibody screen will detect the presence of any unexpected antibodies in patient serum If antibodies are detected, identification should be performed using panel cells (with an autocontrol) Copy for: HAW, SPENCER O. | 3 Major and Minor Crossmatch Why is the minor crossmatch unnecessary? Donated units are tested for antibodies already through antibody paneling. Most blood is transfused as packed cells, having little antibodies The AABB and FDA develop the standards for blood banking; According to the AABB standards: The crossmatch “shall use methods that demonstrate ABO incompatibility and clinically significant antibodies to red cell antigens and shall include an antiglobulin phase” Methods referred by the statement are the IS, Thermophase and AHG phase – in this phases we can achieve demonstrating ABO incompatibility and detect any clinically significant antibodies (IgG) ○ ○ REMEMBER Samples for donor testing must be collected at the same time as the full donor unit Samples must be stored at ref temp for 7 days : The (1) cards, (2) pilot samples and the (3) collecting bags (4) segments must have the SAME UNIQUE NUMBER Segments are a sample of the blood and eliminate having to open the actual unit COMPATIBILITY TESTING ABO/ Rh typing is FIRST performed Antibody screen is performed next If antibodies are NOT DETECTED: Only immediate spin (IS) is performed using patient serum and donor blood suspension This fulfills the AABB standard for ABO incompatibility This is an INCOMPLETE CROSSMATCH ○ Even though incomplete, it is considered to be safe and an effective method ○ However, if there are antibodies detected in the antibody screening, you must perform all the phases in the crossmatching If antibodies ARE DETECTED: Antigen negative units found and X-matched All phases are tested: IS, 37 deg C, AHG This is a COMPLETE CROSSMATCH What we want in cross matching is a non-reactive result (No agglutination) – The units are compatible for transfusion. PROCEDURE SPECIMEN COLLECTION: Donor cells are taken from segments that are attached to the unit itself ○ No need to open the actual bags; this is to prevent contamination of the blood in the bag (easily deteriorates when contaminated) Units of Whole Blood with segments attached Copy for: HAW, SPENCER O. | 4 If no antibody detected = Perform ONLY immediate spin phase If antibody/ previous alloantibody is detected = Perform COMPLETE CROSSMATCH Incompatible Crossmatches CROSSMATCH WILL: Verify donor cell ABO compatibility Detect most antibodies against donor cells CROSSMATCH WILL NOT: Guarantee normal survival of RBCs Prevent patient from developing and antibody Detect all antibodies Prevent delayed transfusion reactions Detect ABO/Rh errors Currently there are no procedures that can guarantee the fate of a unit of blood to be transfused Despite careful in-vitro testing some compatible units may induce hemolytic reaction from the patient Not 100% effective yet, this must not stop us from performing crossmatching properly. Antibody Screen Crossmatch Cause Resolution Pos Neg Antibody directed against antigen on screening cell ID antibody; select antigen negative blood Neg Pos Antibody directed against antigen on donor cell which may not be on screening cell OR donor unit may have IgG previously attached ID antibody + Complete crossmatch; select antigen negative blood OR perform DAT on donor unit Pos Pos Antibodies directed against Antibody ID, select antigen both screening and donor cells negative blood ADDITIONAL INFORMATION ON TYPES OF COMPATIBILITY TESTS Manual (IS and IAT) Gel Technology Electronic (Computerized) Crossmatch Red cell affinity column technology (ReACT) Solid Phase Adherence Assays (SPAA) Copy for: HAW, SPENCER O. | 5 Red Cell Affinity Column Technology – Similar to Gel Tech; the affinity column test cards have an immunoreactive gel and a mixture of the high affinity Fc receptor binding reagent protein G and A, covalently bonded to agarose beads suspended in a viscous solution Solid-Phase Adherence Assay – This can be used to detect BOTH ANTIGEN and ANTIBODY, because this uses antigen-antibody reaction and adherence to a solid phase support system as opposed to hemagglutination in solution to determine if reaction is positive or negative. The card containing the microtubes is then centrifuged at a controlled speed for 10 minutes At the start of centrifugation the cells are separated from the serum; then they meet the AHG contained in microtube Finally the cells are trapped by the gel (if agglutinated) or pellet to the bottom of the tube. CROSSMATCH PHASES PHASE Manual (IS and IAT) PURPOSE DETECTS Detection of IgM; Cold agglutinins Clinically insignificant such as: IS Anti-M Anti-N Anti-Lea Anti-Leb Anti-I Most ABO incompatibilities IgG/ IgM Abs 37°C, LISS This phase is usually read only MACROSCOPICALLY for agglutination Potent Rh Abs IgG Abs AHG Naturally occurring (IgM in nature; developed due to exposure); Acquired (usually of IgG type; either Allo or Autoantibody) IS detect RT reactive antibodies (Auto, Alloantibody, Naturally occurring) IAT detect IgG antibodies (Auto & alloantibody; Acquired) Gel Technology Uses a single type of gel matrix that replaces saline to separate positive and negative reactions – A dextran acrylamide gel that can separate particles based on size is contained in a plastic microtube. Patient serum, and 1% of suspended RBCs in Low Ionic Media (LIM) are dispensed into the microtube and incubated at 37 deg C for 15 minutes This phase is read MICROSCOPICALLY for agglutination Coombs Check Cells (OCC) AHG absence, inadequacy, or neutralization Added to all negative tests and must produce a positive result Rh, Duffy, Kidd, others Sensitized RBCs Limitations of Pretransfusion Testing Hemolytic transfusion reaction, if the patient’s antibody is too weak to be detected. ○ Standard antibody detection methods such as the indirect antiglobulin test require several 100 antibody molecules per red cell to produce detectable reactions A hemolytic transfusion reaction due to patient misidentification. ○ For example: Group A red cells (meant for transfusion to a group O recipient) will be compatible in vitro tests with an incorrect specimen drawn from a Group A person Copy for: HAW, SPENCER O. | 6 Hemolytic transfusion reaction if donor red cells are inadvertently hemolyzed before entering the patient ○ For example: Red cells hemolyzed by an improperly functioning blood warmer or red cells hemolyzed by contact with an ice pack in a transport container. Nonhemolytic transfusion reactions such as allergic, febrile, and other reactions Pretransfusion test are meant to detect ONLY red cell antibodies INCOMPATIBLE CROSSMATCH ABO incompatibility ○ Recheck patient information/ history and blood unit ABO group If possible retype again the blood unit Clinically Significant Antibody ○ DAT and IAT ○ Complete crossmatching phases POST-ANALYTICAL PHASE Involves labeling, inspecting and issuing the blood unit ○ After testing, result is released and the blood unit is also released for transfusion Labeling form includes: Patient’s full name, ID number, ABO/Rh of patient and Unit, Donor #, Compatibility Results, and tech ID Form is attached to the donor unit and only released for the recipient The unit is visually inspected for abnormalities, such as bacterial contamination, clots, etc. ISSUING BLOOD When it’s time to release a blood product to the nurse or physician, a few “checks” must be done 1. Requisition form 2. Comparing requisition form → Donor unit tag → Blood Product Label 3. Name of persons issuing and picking up blood 4. Date and time of release 5. Expiration date What if the unit is unused? Blood CAN BE RETURNED if it is not needed for transfusion Unit closure has to remain unopened; if unused yet it is opened, DISCARD it immediately Storage temperature must have remained in the required range (1°C to 10°C for RBCs) If not at correct temp, unit must be returned within 30 minutes of issue Infusion Device This device acts like a smart IV drip, carefully controlling the exact amount of blood a patient gets at just the right speed. Bacterial Contamination This unit shows bacterial contamination and should NOT be given to the patient (due to it being exposed to the external environment) The plasma in the segments is fine, but the plasma in the unit shows heavy hemolysis from bacteria ○ Hemolysis due to bacteria consuming the red cells Discoloration of blood bags is also a way to detect bacterial contamination. Blood Warmer This device brings the blood product up to near body temperature, preventing shivering and other issues caused by cold blood. Copy for: HAW, SPENCER O. | 7 SPECIAL CIRCUMSTANCES Emergency Release In an emergency (ER or OR), there may not be enough time to test the recipient’s sample In this case, blood is release only when signed by the physician (O negative) The tag must indicate it is “not crossmatched” ○ For the protection of the medical technologist you must include in the note that it was not crossmatched Segments should be retained for crossmatching – in the case that the patient is now available for crossmatching (if pwede na makuhaan ug sample si patient) Every detail is documented (names, dates..) Once the specimen is received, ABO/Rh typing and antibody screening should be performed Crossmatching the segments from the released unit should be done In addition, the lab may crossmatch additional units as a precaution if more blood is needed If death should occur, testing should be complete enough to show that the death was unrelated to an incompatibility The antibodies in the patient serum may no longer be demonstrable because of the dilution of large volumes of plasma and other fluids ○ The tests might miss the patient's own antibodies because all the new blood from the transfusion dilutes them out. ○ So, always check the history of the patient A rapid rise of the antibody titer level and subsequent destruction of donor red cell may occur if antigen positive units is infused ○ If the patient has antibody towards the rbc (antigen) it will cause HTR ○ If time permits always perform antibody screening APPROPRIATE UNITS TO GIVE Patient’s Type 1st Choice Other Choices O O None A A O B B O AB AB A,B,O ABO compatible should always be given first What can be given in an emergency? Group O Rh-Negative red cells or AB plasma ○ Emergency release ○ Women below or of childbearing age ○ Or if in doubt Group O Rh-Positive red cells ○ Used as a substitution ○ Male or Elderly females Massive Transfusion Defined as a transfusion approaching or exceeding the recipient’s own blood volume (about 5 liters or 10-12 units in an adult male) within 24 hour period The original sample no longer represents the patient’s condition ○ The blood in the body of the patient is mostly not his/her original blood anymore but from the transfused blood from other persons Complete crossmatch is not necessary (if no antibodies were detected originally) Give ABO identical units ○ If antibodies were identified, continue to give antigen negative units If the patient is known to have a clinically significant unexpected antibody by checking the history of the patient – all infused units must be tested for and should lack the corresponding antigen if time permits (perform antibody screening) Group O individuals are “UNIVERSAL DONORS”, they can donate blood to any blood group because they have no A or B antigens Grup AB individuals are “UNIVERSAL RECIPIENTS”, they can receive blood from any group because they do not have A or B antibodies. Rh negative blood types are to receive ONLY Rh negative blood units Copy for: HAW, SPENCER O. | 8 Pedipacks are small aliquots of larger units and prepared by the donor facility or hospital lab for infant transfusion. If the selected cells for transfusion are not group O, serum/plasma of the neonate should be tested; we must make sure that the serum has no Anti-A or Anti-B ○ How do we do that? We use A1 cells and B cells Autologous Crossmatching Type & Screen Used to conserve blood inventory On average, a surgical procedure uses about 1 unit of RBCs, however, many times the units are on “hold” in the lab and will not be needed (reducing inventory) For this reason, only a type & screen are performed and if any blood is needed, the sample can be retrieved for crossmatching (only the IS phase is required) ○ This is a way to be prepared for a possible transfusion while being efficient with blood bank resources. If there are antibodies found, the hospital can then take extra steps to find compatible blood for you. If antibodies are identified, then antigen negative blood is reserved or crossmatched Neonatal Transfusion A neonate who is 72 hours Completion of therapy for syphilis/gonorrhea Transfusion of blood, components, human tissue, plasma-derived clotting factor concentrates Human diploid cell-rabies vaccine after animal bite Deferrals Temporary Deferral Recommended if the donor would be eligible at a specific time in the future Prospective donor is unable to donate blood for a limited period of time EXAMPLE: Donor has received a blood transfusion; defer for 12 months from date of transfusion. The donor received vaccination for yellow fever; defer for 2 weeks from date of vaccination. Indefinite Deferral Due to current regulatory requirements that may change in the future Prospective donors are unable to donate blood for someone else for an unspecified period of time due to current regulatory requirements. This donor would not be able to donate blood until the current requirement changes. These donors may be eligible to donate autologous blood. EXAMPLE: Donor states they have lived in England for 1 year in 1989; defer indefinitely Permanent Deferral Based on High-Risk Behavior or a Positive Test Result Prospective donor will NEVER be eligible to donate blood for someone else. Donoors who have had sexual contact with anyone who: Copt for: HAW, SPENCER Y. | 4 1. 2. 3. 4. Has used needles to take drugs NOT prescribed by a physician Has taken clotting factor concentrates for a bleeding problem Has HIV/AIDS or has had a positive test for HIV Women who had sex with man has had sex with other men Conditions for Indefinite/Permanent Deferral History of Viral Hepatitis after the eleventh birthday ○ Hepatitis B is present after eleventh year ○ Blood-borne virus Confirmed positive test for Hepatitis B Surface Antigen (HBsAg) Reactive test to antibodies to Hepatitis B core on more than one occasion Present or past clinical or lab evidence of infection with Hepatitis C Virus, Human T-Lymphotropic Virus, or Human Immunodeficiency Virus History of Babesiosis or Chagas Disease Family history of Creutzfeldt-Jakob Disease Recipient of dura mater or Human Pituitary Growth Hormone Risk of Variant Creutzfeldt-Jakob Disease (vCJD) or Mad Cow Disease ○ Ask if the patient is taking insulin because the insulin source may be from a cow Use of needle to administer nonprescription drugs ○ Drug abuse users Men having sex with men (MSM) since 1977 are Permanently Deferred ○ According to Harmening, patients who are engaging in this activity, as long as the previous sexual contact is 1 year before, and the rest of the requirements are valid ○ For exam purposes, still follow permanently deferred Severe heart or liver disease Deferral Periods for Potential Transfusion-Transmitted Infections Infectious Disease Malaria (Plasmodium spp.) Health History Deferral History of malaria: 3 years Lived in endemic country for 5 consecutive years: 3 years from departure Travel to endemic area: defer for 1 year from departure Babesiosis (Babesia microti) History of Babesiosis: Indefinite deferral Chagas disease (Trypanosoma cruzi) History of Chagas disease: Indefinite deferral Leishmaniasis Travel to Iraq: defer for 1 year from departure Variant Creutzfeldt-Jakob Disease Indefinite deferral by geographic regions: Lived >3 Creutzfeldt-Jakob Diease months in UK from 1980-1996 Lived >5 years in Europe from 1980 to present Family history of Creutzfeldt-Jakob Disease, dura mater transplant, human pituitary derived Growth Hormone: indefinite deferral Deferrals on Medicine Intake Tegison Teratogenic drug Poses problems and complications on neonates PERMANENT DEFFERAL Soriatane Treatment for severe psoriasis 3 YEARS DEFERRAL Proscar 1 MONTH DEFERRAL Treatment for Benign Prostatic Carcinoma 6 MONTHS DEFERRAL Accutane Avodart Hypnotics used at bedtime Blood pressure medications (Free of side effects and CV symptoms) Over-the-counter bronchodilators Decongestants Oral contraceptives Replacement hormones Weight-reduction drugs Mild analgesics Vitamines Tetracyclines and other antibiotics taken for acne DEFERRED: Antibiotics Anticonvulsants Anticoagulants ○ Aspirin → DEFERRED FOR THREE DAYS Insulin (Mad Cow Disease) Antiarrhythmic drugs (heart conditions) PARAMETERS FOR PHYSICAL EXAMINATION Temperature Should not exceed 37.5 degrees Celsius Blood Pressure Systolic and Diastolic blood pressure should be within normal limits and Copt for: HAW, SPENCER Y. | 5 controlled If patient is hypertensive, as long as controlled, it is okay Pulse Therapeutic bleeding WB: No specifications Plasma: 50-100 beats/minute Weight Bag Labelling In-depth discussion on F3 Primary Bag, Satellite Bags Sample tubes Donor registration form 10.5 mL of blood per kilogram of the donor’s weight 110 lb (50 kg) Allogeneic (Regular) Donor Autologous donor Hemoglobin Level 12.5 g/dL 11.0 g/dL Hematocrit Level 38% 33% Qualitative Test for Hgb Determination Copper sulfate If drop of blood floats, 15 minutes: not suitable for the preparation of platelets, fresh frozen plasma, or cryoprecipitate Skin at the venipuncture site must be free of lesions and needle marks. Blood removed for medical purposes such as in polycythemia vera (increased iron blood or hemochromatosis). NOT USED FOR TRANSFUSION Informed Consent Understanding of all the donor Information presented Additional questions Blood tests Notified if testing indicates that the blood presents a risk of transmitting disease Used for other purposes DONOR MUST SIGNATURE AN INFORMED CONSENT Donor Categories Allogeneic “Homologous” and “Random donor” terms used for blood donated by individuals for ANYONE’S use Autologous Donate blood for your own use only MOST SAFE Recipient-specific directed donation Donor called in because blood/blood product is needed for a specific patient Unique, rare blood groups that are only allowed to be donated to specific patients MOST DANGEROUS Other Important details Frequent mixing of blood is needed Volume of blood withdrawn is monitored 2-4 tubes are filled with blood samples before or after the procedure Use of DIVERSION POUCH in units which will be processed for component prep (Platelets) Donor Reactions MILD Syncope (fainting spell) Nausea Hyperventilation Twitching Copt for: HAW, SPENCER Y. | 6 Normovolemic Hemodilution Muscle spasm MODERATE Mild reactions and loss of consciousnesss SEVERE Moderate reactions and convulsions Blood Recovery ADVERSE DONOR REACTIONS Symptom Treatment Weakness, sweating, dizziness, pallor, nausea, vomiting Remove needle and tourniquet; elevate legs above head; apply cold compress from back to the neck Syncope Cold compresses on back of neck Twitching, muscle spasms Have donor cough Hematoma Apply pressure 7-10 minutes; apply ice for 5 minutes Convulsions Call for help; ensure donor’s airways are free Cardiac difficulties Call for emergency help Post-Donation Avoid smoking for 30 mins; avoid alcohol until something has been eaten Drink more fluids for the next 4 hours If dizziness or fainting occurs, lie down or sit with the head between the knees Inform the blood center if any symptoms persist Informed consent should be obtained from patient (donor) No age restriction Amount of blood determined by the weight and size of the patient Hemoglobin: 11 g/dL Hematocrit: 33% Blood collection should be completed 72 hours before the surgery Proper labelling should be followed ABO and D typing should be performed The units cant be crossed over to the general inventory Collection and reinfusion of shed blood Direct Donation Apheresis Whole blood is removed from the donor, component is separated by mechanical means and remainder of blood is returned to the donor Leukapheresis White blood cells are removed Requires drug or sedimenting materials to be given to the donor before collection Plateletpheresis Platelets are removed 48 hours must elapse between donations Donors should not undergo more than twice a week or more than 24 times a year Platelet count: >150,000 uL Plasmapheresis Plasma is removed Donate once every 4 weeks Red Cell Apheresis 2 units of RBCs are removed Performed using cell operator machine (closed system) FDA requirement: donors are larger and have HIGHER Special Blood Collections Preoperative Collection Removing at least 1 or more units of blood at the beginning of the surgery Blood is removed and replaced with a crystalloid or colloid solutions to restore fluid volume Blood is stored for reinfusion during or at the end of the surgery Copt for: HAW, SPENCER Y. | 7 Therapeutic phlebotomy HEMATOCRIT levels Males: 59 kg (130 lbs), 5”1’ Females: 68 kg (150 lbs), 5”5’ HEMATOCRIT: 40% and above Donors deferred for 16 weeks after 2 units RBC donation Withdraw of blood from a patient for medical reasons THERAPEUTIC APHERESIS Blood is removed from the patient Portion that might be contributing to a pathologic condition is retained The remainder is reinfused with a replacement fluid such a colloid or Fresh frozen plasma HBsAg, Anti-HBc, Anti-HCV, NAT (Nucleic Acid Amplification Test) Confirmatory test: Western Blot Additional Tests West nile virus Cytomegalovirus Chagas Disease Malaria Babesiosis Prion Disease Bacterial Contamination Most blood units at risk: PLATELETS (Normal skin flora) Most common reactions: Fever, shock, DIC ○ Transfusion Service Testing Testing of Donor’s Blood Two Categories Immunohematologic Testings ABO and D Typing Antibody Screen ABO and D typing Antibody screen Infectious Disease Screening Syphilis Hepatitis virus HIV and others Methods: Microplate, Solid Phase Adherence, Gel Test Any ABO and D typing discrepancies should be resolved before labelling the donor unit Detects unexpected antibodies in the donor’s plasma Most important antibodies detected are those produced after transfusion or pregnancy If clinically significant antibodies are present ○ PLASMA AND PLATELETS are not used for transfusion ○ If used, antibody interpretation is required on the label of the component Serologic Test for Syphilis Methods: Rapid plasma reagin Hemagglutination test Confirmatory test: Fluorescent treponemal antibody-absorption Viral Hepatitis Quarantine and Recipient Tracing (Look-Back) The only repeat testing required is ABO on red cell products D typing (IS) on D negative red cell products Plasma products (Fresh frozen plasma, Cryoprecipitate, Platelets) do not require any testing Donor samples must be stored at 1-6 degrees Celsius at least 7 days after transfusion ADSOL unit transfused today must save sprig for one week Many facilities will pull a sprig from each donor during processing and save all sprigs for 49 days, regardless of expiration of unit Quarantine of prior collections from the donor Notification of facilities (eg hospitals) that receive products to quarantine prior collections Further testing of the donor Notification of recipients A, B, C, D, E, G ○ Check Mycology and Virology F1 trans Four current hepatitis tests Copt for: HAW, SPENCER Y. | 8 [MLS 416-LEC] Immunohematology - Lecture F3: Blood Component Preparation and Therapy Professor: Bea Angelli Laude, RMT Date: May 1, 2024 INTRODUCTION Blood and its components are utilized NOW as part of the treatment regimen of patients, in what we call as the process of HEMOTHERAPY Each unit of whole blood can be separated into different components, depending on what is necessary to be transfused to the patient This process of blood preparation is going to MAXIMIZE the usage of blood in which component transfusion is going to be much more safer than the use of WHOLE BLOOD Goals of Blood Collection Maintain viability and function ○ Once the blood is being transfused to the patient, it is still going to function and aid in the recovery process of the patient. Prevent physical changes ○ We have to ensure that we have added additive solutions & anticoagulants in order to PREVENT further physical changes that have occured on the blood component upon storage Minimize bacterial contamination ○ Less occurrence of transfusion-associated infections → over time, there were a lot of blood transfusion procedures performed and were considered to be unsuccessful Thanks to the advancement of technology (Discovery of plastics vs. glass) These help in the storage life of blood components Equipment Manufacturing Quality Control Blood bags → collect & store blood components ○ Anticoagulants and additives present Filtration → ensures no contamination from one component to another Centrifuge → most important - made component separation possible → Allows MedTechs to follow CGMP (Current Good Manufacturing Practices) Collection of methods used in the facility/controls used for the manufacture, processing, packing, or holding of a blood product to ENSURE that it meets the safety, purity, and potency standard. Storage Freezers Refrigerators Plated agitator machines Documentation Capture the critical steps in the blood manufacturing process: Personnel Equipment used Date and time → These records will be retained and must be protected from destruction as these are REQUIRED by the Department of Health (DOH) during inspection. All blood banks will be under DOH US Blood Banks are under FDA and AABB Other countries have the BECS (Blood Establishment Computer System) ○ Able to identify blood that are not tested with infectious diseases ○ Able to identify blood required to be quarantined → considered to be UNACCEPTABLE blood components → Philippines : perform manual recording, some are encoded in the computer systems → Other countries : have barcodes Blood Collection and Storage Anticoagulants are responsible for preventing clotting process; maintains blood in fluid state Additive solutions → allows us to store blood on a specific number of days depending on the type of solution that is contained in the blood bag → Must be maintained in a closed system Ensures that we have maintained sterility Recall that we should minimize bacterial contamination Once exposed to air (in the tubing, blood bag itself, top/bottom, entry of air): now considered to be an open-system ○ Example: Transfer of blood component in a particular blood bag to another blood bag ○ ALLOWABLE STORAGE TIME FOR OPEN SYSTEM → reduced Why? Because bacteria might contaminate it → discard! Copy for: HAW, SPENCER Y. | 1 Single Bag ONLY WHOLE BLOOD will be collected (cannot be separated into diff components) Double Bag Plasma and packed red cells Quadruple bags Packed RBCs, platelet concentrate, plasma, cyroprecipitate Anticoagulant Preservative Solutions and Storage Lesions FDA (Food and Drug Administration) : Storage limits and temperature criteria for preservative solutions ○ able to support at least 75% of original RBCs in the recipient's circulation for atleast 24 hours after transfusion, with less than 1% hemolysis STORAGE LESIONS – all biochemical changes that occur in the blood upon storage (collectively known as) INCREASED Triple bags Packed RBCs, platelet concentrate, plasma Hemoglobin Freed when there is hemolysis Potassium Released Visible cells Cells present in the blood bank could be senescent or newly matured cells Some of them are going to be destroyed during storage, some will stay (can survive for 120 days) DECREASED pH Becomes acidic due to the buildup of lactic acid as a byproduct of glucose metabolism in order to produce the energy utilized by the red cells Sodium Possibly from gradual breakdown of rbcs during storage and eventual release of cellular contents that includes sodium Copy for: HAW, SPENCER Y. | 2 ATP 2,3-DPG Red blood cells continue to use ATP for various cellular functions even during storage. However, unlike in the body, stored blood lacks a constant supply of oxygen and glucose, which are essential for ATP production. This limited supply leads to a depletion of ATP stores within the RBCs. Cells are going to be inactive and there will be depletion of the glucose sources Additive solutions tend to contain glucose that would enable the cell to survive for a particular number of days, depending on the preservative solution incorporated in the blood bank Necessary for the oxygen release of the red cells ○ Hgb has high affinity to oxygen → OXYGEN cannot be released MOST IMPORTANT BIOCHEMICAL CHANGE – DECREASED 2,3-DPG, DECREASED OXYGEN RELEASE ○ In newborns undergoing exchange transfusion, it is necessary to make use of FRESH BLOOD (fresh whole blood or freshly separated packed RBCS from whole blood) ○ This is to ensure 2,3-BPG is not entirely separated in which it is in enough amount to serve its function for oxygen release For adults, the doctor may require it – but for newborns it is required to have enough amounts of 2,3-DPG present on the blodo component plasma – in order to generate ATP, once taken out of the body and separated from the plasma, its glucose source is going to be depleted Adenine Acts as substrate for red cell ATP synthesis Citrate Prevents coagulation by chelating calcium Protects the red cell membrane from hemolysis Sodium biphosphate Prevents excessive decrease in pH Acts as a buffer Remember: one of storage lesions is the drop of pH due to the accumulation of lactic acid ○ Red cells are also going to be destroyed if the environment is going to be very acidic Mannitol Osmotic diuretic acts as a RBC membrane stabilizer to avoid hemolysis Anticogulant/Preservative Solutions incorporated in blood bags Anticoagulant/Preservative Preservative Solutions Chemical Dextrose Purpose Source of sugar so red cells [are] Support[ed] with ATP generation via glycolytic pathway Remember: ATP is its main source of energy Recall: Red cells are dependent on the glucose content of Number of Days ACD (Acid Citrate Dextrose) Rarely used because of the acid (should not react w/ lactic acid; makes red cells more prone to hemolysis) 21 CPD (Citrate-Phosphate-Dextrose) 21 CP2D (Citrate-Phosphate-Double-Dextrose) 21 CPDA1 (Citrate-Phosphate-Dextrose-Adenine) 35 AS- 1/AS-5 (dextrose, adenine, mannitol, saline) Mannitol as the membrane stabilizer 42 AS-7 (dextrose, adenine, mannitol, saline) 42 AS- 3 (dextrose, adenine, saline, citrate, phosphate) Citrate and Phosphate are the membrane stabilizers 42 Additive solutions are added to the red cells/packed red cells in order to add more nutrients so it could extend its shelf life Copy for: HAW, SPENCER Y. | 3 Despite its disadvantage, it is valuable for preserving autologous units and red cells with the rare phenotypes Blood Component Preparation → a lot of factors can affect blood component preparation, including temperature and time constraint Specific components of blood are affected Remember that components are going to be separate from whole blood Prior to separation, ○ → If we are to add additive solutions in the packed RBC, more plasma can be removed mainly because the additive solution is ADDED to maintain red cell metabolism during storage → Normally if there is no additive solution, a small amount of plasma will be retained in the packed red cell so it is going to act as a diluent and additional glucose source apart from the preservative solution → MORE PLASMA will be extracted because the additive solution will act as diluent, and provide more sugar source in order to produce ADP → will minimize storage hemolysis to