Tm14 - red cell typing methods

Red Cell Typing Methods
Method 2.1. Slide Test for
with the manufacturer’s instructions.
Determination of ABO Type
of Red Cells
Procedure
Principle
Place 1 drop of anti-A,B on a third slide, Specimen
if parallel tests are to be performed withthis reagent, or on a single clean, labeled The reagent manufacturer’s instructions must slide if this is the only test performed.
be consulted before slide tests are performed; slide tests with whole blood, while others spec- ify the use of red cell suspensions of lighter cells to be tested. (Consult reagent man- concentrations prepared in saline, serum, or ufacturer’s instructions to determine the Reagents
oughly, using a clean applicator stick for Copyright 2002 by the AABB. All rights reserved.
AABB Technical Manual
Gently tilt the slide continuously for up men requirements. Generally, clotted or anti- to 2 minutes. Do not place the slide over ABO testing. The red cells may be suspended in native serum, plasma, or saline, or may be Read, interpret, and record the results of Reagents
Interpretation
Anti-A,B. Note: Use of this reagent is op- A , A , and B red cells. These can be ob- end of 2 minutes is a negative result.
suspension on each day of use. (Note:The use of A cells is optional.) with the manufacturer’s instructions.
of contact between specimen and tech-nologist than occurs with tube testing.
Procedures
detailed in the facility’s procedures man- Place 1 drop of anti-A in a clean, labeled Slide testing is not suitable for detection Place 1 drop of anti-B in a clean, labeled ABO serum groups cannot be accurately Add to each tube 1 drop of a 2% to 5%suspension (in saline, serum, or plasma) Method 2.2. Tube Tests for
Determination of ABO Type
tively, the equivalent amount of red cellscan be transferred to each tube with of Red Cells and Serum
Principle
Mix the contents of the tubes gentlyand centrifuge them according to ples of testing for ABO types. The following Typically, this is 15 to 30 seconds at ap- procedure is an acceptable representative proximately 900 to 1000 × g.
method, but the manufacturer’s instructions for the specific reagents must be consulted.
Read, interpret, and record test results.
Specimen
The reagent manufacturer’s package insert must be consulted to determine specific speci- Copyright 2002 by the AABB. All rights reserved.
Methods 2: Red Cell Typing Methods
to 15 minutes to enhance weak reactions. Seefor discussion of weakly reactive Label two clean test tubes as A and B.
(Note: Label additional tube if optionaltest with A red cells is to be performed.) Add 2 or 3 drops of serum to each tube.
Method 2.3. Microplate Test
for Determination of ABO
Add 1 drop of B reagent cells to the tubelabeled B.
Type of Red Cells and Serum
this optional test is being performed.
Principle
ples of testing for ABO. Microplate techniques manufacturer’s directions. Typically, this can be used to test for antigens on red cells A microplate can be considered as a matrix of 96 “short” test tubes; the principles that ap- ply to hemagglutination in tube tests also ap- Microplates may be rigid or flexible, with Read, interpret, and record test results.
either U-shaped or V-shaped bottoms.
obtained in testing red cells (see above).
cause results can be read either after centri-fuging the plate and observing the characteris- Interpretation
tics of resuspended red cells or by observing Agglutination of tested red cells and ei- the streaming pattern of the cells when the plate is placed at an angle. Either reading tech- on serum constitute positive test results.
nique permits estimation of the strength of ag-glutination.
A smooth cell suspension after resus-pension of the cell button is a negative Specimen
Interpretation of serum and cell tests for Equipment
the patient’s or donor’s ABO type (see to a row of wells. Special plate carriers can be purchased to fit common table-top centrifuges.
Positive reactions characteristically show 3+ to 4+ agglutination by reagent ABO antibod- ies; reactions between test serum and reagent red cells are often weaker. The serum tests between positive and negative tests.
may be incubated at room temperature for 5 Copyright 2002 by the AABB. All rights reserved.
AABB Technical Manual
400 × g for 30 seconds after addi- results interfaced with a microprocessor.
The automated reader passes lightbeams through the bottoms of the Reagents
ing reagents that are licensed by the Food and Drug Administration (FDA) for use as undi- luted reagents in microplate tests. Most re- agent red cells licensed for use in tube tests can be used in U-bottom plates without addi- tional preparation other than dilution to a 2% reactions and prints the blood testing re- to 5% concentration. If an FDA-licensed labo- ratory wishes to employ reagents in a manner not specified by the manufacturer (eg, diluting and cell specimens must be followed.
an antiserum before use), the laboratory must submit a description of its procedure to the Center for Biologics Evaluation and Research (CBER). The user who changes the test condi- forces, expressed as g, are suggested.
tions assumes responsibility for appropriate reagent evaluation. Unlicensed blood banks and transfusion services do not need FDA ap- If the antisera directions require testing proval for microplate use, but must have docu- cedure that differs from the manufacturer’s in- red cells because the cell samples resus- Anti-A,B. Note: Use of this reagent is op- 700 × g for 5 seconds for red cell 700 × g for 20 seconds for wash-ing red cells for the antihuman Procedures
700 × g for 5 seconds after addi- Place 1 drop of anti-A and anti-B in sep- 400 × g for 30 seconds for red cell 400 × g for 3 minutes for washing pension of red cells to each well contain- Copyright 2002 by the AABB. All rights reserved.
Methods 2: Red Cell Typing Methods
conditions established for the centrifuge.
fore an interpretation is recorded for the Read, interpret, and record results. Com-pare test results on red cells with thoseobtained in testing serum.
Method 2.4. Confirmation of
Weak A or B Subgroup by
Adsorption and Elution
Add 1 drop of a 2% to 5% suspension ofreagent A and B red cells to separate Principle
clean wells of a U-bottom microplate.
(Note: If optional test on A cells will be Add 1 drop of serum or plasma undertest to each well.
Specimen
Reagents
conditions established for the centrifuge.
tapping the plate or with the aid of a me- Read, interpret, and record results. Com- pare test results on serum with those ob- Procedure
To enhance weak serum reactions, the plates may be incubated at room temperature for 5 to 10 minutes, and then the centrifugation, reading, and recording steps may be repeated.
Add 1 mL of reagent anti-A (if a weakvariant of A is suspected) or 1 mL of Interpretation
Agglutination in any well of red cell tests resuspension of the cell button is a nega- cells. Remove all supernatant reagent.
Transfer the red cells to a clean test tube.
Copyright 2002 by the AABB. All rights reserved.
AABB Technical Manual
(4 C) saline. Save an aliquot of the final If the wash solution reacts with the A or wash supernatant fluid and test it in par- B cells, tests on the eluate cannot be con- Centrifuge to pack the cells and transfer negative controls and tested in parallel.
Reference
tion (from step 6), in parallel, againstthree examples of group O cells and Beattie KM. Identifying the causes of weak or “missing”antigens in ABO grouping tests. In: The investigation of typing and compatibility problems caused by red blood cells. Washington, DC: American Association of Blood anti-A, B cells for anti-B). Add 2 drops ofeluate or wash to 1 drop of cells, and ex-amine for agglutination after immediatecentrifugation; if negative, incubate 15 Method 2.5. Saliva Testing
to 30 minutes at room temperature. Ifthese phases are both negative a 15- for A, B, H, Lea, and Leb
minute incubation at 37 C and the indi-rect antiglobulin test may also be per- Principle
Approximately 78% of individuals possess the Interpretation
Se gene that governs the secretion of water-soluble ABH antigens into all body fluids with the exception of cerebrospinal fluid. These se- eluate, hence the presence of A or B anti- creted antigens can be demonstrated in saliva gen on the test cells, is confirmed if: a) the by inhibition tests with ABH and Lewis anti- positive cells, at any phase; b) the eluateis nonreactive at all phases with all three Specimen
group O cells; and c) the final wash solu-tion is nonreactive with all six cells.
or B cells, it may indicate that the test cells do not express the antigen and can- the A or B cells and also with some or all Centrifuge saliva at 900 to 1000 × g for Copyright 2002 by the AABB. All rights reserved.
Methods 2: Red Cell Typing Methods
Transfer supernatant to a clean test tube status. Use Le(a+b–) red cells to deter- and place in boiling waterbath for 8 to 10 minutes to inactivate salivary enzymes.
Recentrifuge at 900 to 1000 × g for 8 to card the opaque or semisolid material.
Dilute the supernatant fluid with an within several hours. If testing will not be labeled “Secretor,” “Nonsecretor,” samples retain activity for several years.
“Saline,” and “Unknown.” For Lewisstudies, they will be “Lewis-positive,” Reagents
“Lewis-negative,” “Saline,” and “Un- Human (polyclonal) anti-A and anti-B.
not be appropriate for use; therefore, ap- the “Secretor,” “Nonsecretor,” and “Un- Anti-H lectin from Ulex europaeus. Ob- line extraction of Ulex europaeus seeds.
Polyclonal (rabbit/goat/human) anti-Lea.
ability of monoclonal Lewis antibodies.
Group O, Le(a+b–) red cells, as used for antibody detection or identification (see nonsecretors, to use as positive and neg- Procedures
Interpretation
Agglutination of indicator cells by anti- body in tubes containing saliva indicates that the saliva does not contain the cor- indicator cells after incubation with sa- red cells. Use A , B, or O cells to deter- liva indicates that the saliva contains the Copyright 2002 by the AABB. All rights reserved.
AABB Technical Manual
Failure of antibody in the saline control line dilutions of saliva. The higher the di- tube to agglutinate indicator cells invali- dates the results of saliva tests; this usu- tivity, the more blood group substance is ally reflects use of reagents that are too present in the saliva. Saliva should be di- with diluted anti-A and anti-B reagents.
The appropriate dilution of anti-A or tus, use saliva from previously tested Se to have Leb as well as Lea in the saliva. A control; use saliva from a Le(a–b–) per-son as the negative control. Aliquots of B, or H substances lacks the Se gene and blood group activity by testing serial sa- Table 2.5-1. Interpretation of Saliva Testing
Testing with Anti-H
Se Saliva
Non-Se Saliva
(H Substance
(H Substance Not
Present)
Present)
(Dilution Control)
Interpretation
Testing With Anti-Lea
Le-negative
Le-positive Saliva
(Dilution Control)
Interpretation
*A Lewis-positive person shown to be a secretor of ABH can be assumed to have Leb as well as Lea in saliva. A Le(a+) personwho is sese and does not secrete ABH substance will have only Lea in saliva.
Copyright 2002 by the AABB. All rights reserved.
Methods 2: Red Cell Typing Methods
Method 2.6. Slide Test for
may cause formation of rouleaux, whichmay be mistaken for agglutination.
Determination of Rh Type
Interpret and record the results of the re- Principle
Interpretation
Specimen
tute a positive test result and indicatethat the cells being tested are D positive.
Reagents
No agglutination with either anti-D orthe Rh control suggests that the cells are that are not agglutinated on slide testing.
slide, results of the anti-D test must not Rh control reagent: The manufacturer’sinstructions will indicate the type of re- mixture must not be confused with ag-glutination.
Procedure
Place 1 drop of anti-D serum onto aclean, labeled slide.
reagent onto a second labeled slide.
nologist than occurs with tube testing.
(The manufacturer’s instructions will in- detailed in the facility’s procedures man- For slide tests using low-protein anti-D, a negative result on slide testing with ei- or plasma) of the red cells to be tested.
reagent, using a clean applicator stick foreach test, spreading the reaction mixtureover an area approximately 20 mm ×40 mm.
Method 2.7. Tube Test for
Place both slides on the viewbox and tilt Determination of Rh Type
them gently and continuously to ob-serve them for agglutination (see note Principle
1). Most manufacturers stipulate that thetest must be read within 2 minutes be- Copyright 2002 by the AABB. All rights reserved.
AABB Technical Manual
Specimen
point. Donor blood must be furthertested for the presence of weak D anti- Reagents
steps 1-5, above, may be used to test forweak D, providing the manufacturer’s Reagent anti-D: Suitable reagents include directions state that the reagent is suit- (eg, monoclonal) reagents. Follow the in-structions from the manufacturer of the anti-D in use before performing tubetests. The method presented here is a rep- instructions will indicate the type of con- choose to do additional testing on results with agglutination of <2+. Requiredtesting must be defined in the facility’s Procedure
low-protein anti-D reagent has been used.
Place 1 drop of the appropriate controlreagent in a second labeled tube.
Add to each tube 1 drop of a 2% to 5%suspension (in saline, serum or plasma) Method 2.8. Microplate Test
for Determination of Rh Type
tively, the equivalent amount of red cellscan be transferred to each tube with Principle
and speed specified by the manufacturer.
Gently resuspend the cell button and ex-amine for agglutination. If a stick was Specimen
used to transfer the red cells, adding 1drop of saline to each tube will make it easier to resuspend the cell button.
coagulated samples may be used for Rh test-ing. Follow the manufacturer’s instructions for Interpretation
Reagents
Agglutination in the anti-D tube, com-bined with a smooth suspension in the Use only anti-D approved for use in micro- control tube, indicates that the red cells Procedure
the anti-D and the control tubes is a neg- The following is a representative method; the ative test result. Specimens from patients manufacturer’s instructions should be fol- lowed for specific reagents and equipment.
Copyright 2002 by the AABB. All rights reserved.
Methods 2: Red Cell Typing Methods
Place 1 drop of the Rh reagent in a clean Specimen
well of the microplate. If the reagent re- quires the use of an Rh control, add 1drop of the control to a second well.
Reagents
3. Mix the contents of the wells by gently 4. Centrifuge the plate at appropriate con- ditions established for the centrifuge.
5. Resuspend the cell buttons manually by reagent is used for this purpose. Not ev- tapping the plate or with the aid of a me- 6. Read, interpret, and record results.
8. Centrifuge the plate at appropriate con- ditions established for the centrifuge.
9. Resuspend the cell buttons manually by tapping the plate or with the aid of a me- Read, interpret, and record results.
Procedure
Interpretation
If the original, direct test with anti-D was per- Agglutination with Rh reagent after the imme- formed by tube testing, the same tube may be diate-spin or 37 C incubation phase indicates used for the weak D test, providing the manu- a positive test provided there is no agglutina- facturer’s directions so state. In this case, pro- ceed directly to step 4, after recording the for determining Rh phenotypes from reactions original anti-D tube test as negative.
obtained with Rh blood typing reagents.
Place 1 drop of anti-D serum in a clean,labeled test tube.
Place 1 drop of the appropriate controlreagent in a second labeled test tube.
Refer to the manufacturer’s instructions for suspension in saline of the red cells to betested. It is permissible to use a directantiglobulin test on the test cells as a Method 2.9. Test for Weak D
control, but an indirect antiglobulin pro-cedure with Rh control reagent is prefer- Principle
weakly that most anti-D reagents do not di- rectly agglutinate the cells. Weak D expression can be recognized most reliably by an indirect to reagent manufacturer’s directions.
antiglobulin procedure after incubation of the Copyright 2002 by the AABB. All rights reserved.
AABB Technical Manual
5. If a reading is desired after the 37 C to the reagent manufacturer’s directions.
given until the D type can be resolved. If test red cells are strongly agglutinated in negative antiglobulin tests are discussed tube, record the test sample as D positive and do not proceed with the antiglobulinphase of the test.
If the test cells are not agglutinated or re-sults are doubtful, wash the cells three or Some facilities may elect to do an additional four times with large volumes of saline.
reading after the 37 C incubation before com- 8. After the final wash, decant the saline pleting the antiglobulin phase of testing. Refer to the manufacturer’s instructions. If this op- dry, and add 1 or 2 drops of antiglobulin tional reading is performed, the facility’s pro- reagent, according to the manufacturer’s cedures manual should indicate its policy on interpretation of this result and on additionaltesting requirements.
9. Mix gently and centrifuge according to Gently resuspend each cell button, ex-amine them for agglutination, and gradeand record the test result.
Method 2.10. Preparation
If the test result is negative, add known and Use of Lectins
IgG-sensitized red cells, and repeat cen-trifugation and examination for aggluti-nation. Agglutination at this point Principle
Saline extracts of seeds react with specific car- globulin reagent in the test mixture.
bohydrates on cell membranes and make use-ful typing reagents that are highly specific at Interpretation
appropriate dilutions. Diluted extract of Doli- chos biflorus agglutinates A red cells but not antiglobulin test must accompany the test A . Ulex europaeus extract reacts with the H determinant; it agglutinates in a manner pro- tube and none in the control tube consti- tutes a positive test result. The blood must (O>A >B>A B>A >A B red cells). Other be classified as D positive. It is incorrect to lectins useful for special purposes include report such red cells as being “D negative, Arachis hypogaea (anti-T), Glycine max (anti-T, -Tn), Vicia graminea (anti-N), and the Salvia lectins (S. horminum, anti-Tn/Cad; S. sclarea, with anti-D is a negative result, indicat- anti-Tn). To investigate red cell polyagglutina- tion, prepare and test the cells with Arachis, Glycine, Salvia, and Dolichos lectins. The antic- If there is agglutination at any phase in ipated reactions with various types of poly- the control tube, no valid interpretation Copyright 2002 by the AABB. All rights reserved.
Methods 2: Red Cell Typing Methods
Table 2.10-1. Reactions Between
Lectins and Polyagglutinable Red
The lectin should agglutinate A1and A B cells but not A , A B, B, or agent purposes, add enough salineto the extract so there is 3+ or 4+agglutination of A and A B cells, Reagents
Seeds: Arachis hypogaea (peanuts) and Glycine max (soy beans) may be obtained from health-food stores, pharmacies, or commercial seed companies. The seeds should be raw.
Procedure
in the order of O>A >B> A >A B.
In a large test tube or small beaker, place A or A B cells are not agglutinated.
their volume of saline. (Seeds vary in thequantity of saline they absorb.) 12 hours, stirring or inverting occasion- to obtain clear supernatant. Collect, and then filter the supernatant fluid and dis- Determine activity of extract with appro- Tests should include a positive and nega- Copyright 2002 by the AABB. All rights reserved.
AABB Technical Manual
Method 2.11. Use of
Method 2.12. Gentle Heat
Sulfhydryl Reagents to
Elution for Testing Red Cells
Disperse Autoagglutination
with a Positive DAT
Principle
Principle
When red cells are heavily coated with IgG, testing with antiglobulin-reactive sera is diffi-cult and testing with high-protein agglutinat- Specimen
ing reagents is impractical. To perform red cell antigen typing, it may be necessary to dissoci-ate antibody from the cells by elution without Reagents
damaging membrane integrity or alteringantigen expression. The gentle heat elution procedure employed to prepare immunoglob- ulin-free red cells differs from procedures in- phate-buffered saline (PBS) at pH 7.3; or 0.1 M 2-mercaptoethanol (2-ME), 0.7mL of stock solution of 14 M 2-ME di- Phosphate-buffered saline at pH 7.3.
Procedure
Specimen
Test cells with a positive direct antiglobulin Add an equal quantity of 0.01 M DTT inPBS, or 0.1 M 2-ME in PBS, to the cells.
Procedure
of normal saline in a test tube of appro- cells positive for the antigen under test.
This will provide a check that the elutiontechnique does not destroy the antigen This procedure is normally used only for ABO forward grouping, Rh testing, and the direct antiglobulin test. Some antigens, in particular Jsa and Jsb , may be weakened or destroyed by be roughly proportional to the degree ofantibody Reference
strength of antiglobulin reactivity.
Reid M E. Autoagglutination dispersal utilizingsulfhydryl compounds. Transfusion 1978;18:353-5.
Copyright 2002 by the AABB. All rights reserved.
Methods 2: Red Cell Typing Methods
Test the person’s cells for degree of anti- Specimen
Red cells with a positive DAT due to IgG coat- antiglobulin test on the treated cells with the antiglobulin results on untreated redcells. If the antibody coating is reduced Reagents
similarly be subjected to a second treat- Test the treated cells for the desired anti- Control red cells, carrying a single-doseexpression of antigens for which the test IgM monoclonal reagents are available;these reagents cause direct agglutination Procedure
As with untreated patient cells, results of solution. Similarly treat the control sam- antigen testing in recently transfused pa- tients should be interpreted with caution Remove a small aliquot (eg, 1 drop) ofthe treated test cells and wash them fourtimes with saline.
Test the washed cells with anti-IgG.
Method 2.13. Dissociation of
IgG by Chloroquine for Red
nonreactive with anti-IgG, wash the totalvolumes of treated test cells and control Cell Antigen Testing of Red
Cells with a Positive DAT
2% to 5% suspension in solution to usein subsequent blood typing tests.
If the treated red cells still react with Principle
Red cells with a positive direct antiglobulin test (DAT) cannot be tested accurately with blood typing reagents that require an indirect antiglobulin technique. Under controlled con- riod of 2 hours), until the sample tested ditions, chloroquine diphosphate dissociates IgG from the red cell membrane with little or no damage to its integrity. Use of this proce-dure permits complete phenotyping of red cells coated with warm-reactive autoantibody, including tests with reagents solely reactive by Copyright 2002 by the AABB. All rights reserved.
AABB Technical Manual
This procedure is routinely used for blood typing tests or adsorption procedures. All common red cell antigens can be detected af- ter treatment with acid glycine/EDTA except antigens of the Kell system and Er antigens.
Thus cells treated in this manner cannot be cause hemolysis and loss of red cell anti-gens.
Specimen
Reagents
control cells for each antigen tested.
strongly positive initial test, may only be turer’s instructions should be followed References
Procedure
Edwards JM, Moulds JJ, Judd WJ. Chloroquine Wash the red cells to be treated six times diphosphate dissociation of antigen-antibody com- plexes: A new technique for phenotyping rbcs witha positive direct antiglobulin test. Transfusion 2. In a test tube, mix together 20 volumes Swanson JL, Sastamoinen R. Chloroquine strippingof the HLA-A,B antigens from red cells (letter).
3. Place 10 volumes of washed red cells in Method 2.14. Acid
4. Add 20 volumes of acid glycine/EDTA.
Glycine/EDTA Method to
5. Mix the contents of the tube thoroughly.
6. Incubate the mixture at room tempera- Remove Antibodies from Red
ture for no more than 2 to 3 minutes.
Add one volume of 1.0 M TRIS-NaCland mix the contents of the tube.
Principle
8. Centrifuge at 900 to 1000 × g for 1 to 2 Acid glycine/EDTA can be used to dissociate antibody molecules from red cell membranes.
Copyright 2002 by the AABB. All rights reserved.
Methods 2: Red Cell Typing Methods
9. Wash the red cells four times with saline.
ing a simple method for recovering autologous cells in a blood sample from recently transfused patients. Note: Red cells from patients with he- moglobin S or spherocytic disorders are not ef- tion procedures. If the DAT is still a posi- Specimen
Red cells from whole blood collected into glycine/EDTA causes irreversible dam-age to cell membranes.
Materials
Procedure
turer’s instructions should be followedfor testing and controls.
Wash the red cells three times in saline.
For the last wash, centrifuge them at References
900 to 1000 g for 5 to 15 minutes. Re- Louie JE, Jiang AF, Zaroulis CG. Preparation of in- tact antibody-free red cells in autoimmune hemolytic anemia (abstract). Transfusion1986;26:550.
Champagne K, Spruell P, Chen J, et al. EDTA/ glycine-acid vs. chloroquine diphosphate treat-ment for stripping Bg antigens from red blood cells (abstract). Transfusion 1996;36(Suppl):21S.
Reid ME, Lomas-Francis C. The blood group anti-gen factsbook. New York: Academic Press, Centrifuge all tubes in a microhemato-crit centrifuge for 15 minutes.
Method 2.15. Separation of
Transfused from Autologous
low the top of the column of red cells.
This 5-mm segment contains the least Red Cells by Simple
Centrifugation
Principle
Newly formed autologous red cells generally add saline, and mix well to flush the red have a lower specific gravity than transfused cells from the microhematocrit tubes.
red cells and may be separated from the trans- fused population by simple centrifugation.
× g for 1 minute and remove the empty Newly formed autologous cells concentrate at hematocrit tubes, or (b) transfer the sa- the top of the column of red cells when blood is centrifuged in a microhematocrit tube, provid- Copyright 2002 by the AABB. All rights reserved.
AABB Technical Manual
lysis by hypotonic saline, in contrast to red cells from normal persons and those with he- moglobin S trait. This procedure permits isola-tion of autologous red cells from patients with have elapsed than if the sample has beenobtained shortly after transfusion.
Specimen
ously while the microhematocrit tubesare being filled.
Reagents
above-normal numbers of reticulocytes.
tients with inadequate reticulocyte pro-duction.
Procedure
Some red cell antigens may not be asstrongly expressed on reticulocytes as on Place 4 or 5 drops of red cells into a 10 be given to determinations of the E, e, c, or until the supernatant fluid no longercontains grossly visible hemoglobin. For References
each wash, centrifuge at 1000 × g for 1 Reid ME, Toy P. Simplified method for recovery of autologous red blood cells from transfused pa- tients. Am J Clin Pathol 1983;79:364-6.
Vengelen-Tyler V, Gonzales B. Reticulocyte rich RBCs will give weak reactions with many blood typ- fuge at 200 × g for 2 minutes to facili- ing antisera (abstract). Transfusion 1985;25: 476.
Resuspend the remaining intact red cellsto a 2% to 5% concentration for pheno- Method 2.16. Separation of
Transfused Red Cells from
Autologous Red Cells in
Patients with Hemoglobin S
Larger volumes, for use in adsorption studies,can be processed in a 16 × 100-mm test tube.
Disease
Principle
Reference
Red cells from patients with sickle cell disease, Brown D. A rapid method for harvesting autologous redcells from patients with hemoglobin S disease. Transfu- either hemoglobin SS or SC, are resistant to Copyright 2002 by the AABB. All rights reserved.

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