Vox Sang. 46: 286-290 (1984) Q IYX4 S. Karger AG. Basel 0042-9007/84/0465-0286 $2.75/0

Normal Survival of Rho (D) Negative, LW (a+) Red Cells in a Patient with Allo-Anti LW"

Elizabeth Cummings", Patricia Pisciottd', Gerald Rothh aConnecticut Red Cross; Department of Laboratory Medicine, University of Connecticut Health Center, Farmington, Conn., USA

Abstract. The serum ofan Rho (D)+ Caucasian female with a prior history of transfusions contained anti-LW a, reacting moderately with Rho (D)+ and weakly with Rho (D)- red cells at the antiglobulin phase. Since transfusions were required, Chromium (5'Cr) survival studies and mononuclear phagocyte assays (MPA) were used to predict in vivo survival. The MPA value of 6% of the positive control predicted a low likelihood of clinically significant extravascular destruction and 51Cr survival studies indicated greater than 95% survival at 1 h and 78% survival at 20 h for rr erythrocytes. Transfusion of4 units of Rho (D)-, serologically incompatible red cells increased the hemoglobin from 5.2 to 11.4 g/dl and the titer of anti- LW" against rr red cells from 4 to 1,024 7 days after the transfusion. A second 5'Cr survival also indicated normal survival of rr erythrocytes. 3 additional Rho (D)- units were success- fully transfused.

Introduction

One serious problem confronting blood bankers is finding serologically compatible blood for patients who have an antibody directed against a high incidence antigen. Since limited information is available con- cerning the clinical significance of many of these antibodies, additional tests, such as the mononuclear phagocyte assay (MPA) [4, 5, 81 and Chromium (51Cr) survival studies [3,6], have been utilized to assist in predict- ing in vivo survival.

Successful transfusion of over 50 units of LW positive blood to a patient with anti-LW has been previously reported [9]. In that case, prior to transfusion the patient's red cells had positive direct antiglobulin test and anti-LW could be demonstrated in the eluate. Anti-LW was also present in the se- rum. The patient's red cells, collected during the course of transfusion therapy, typed as LW negative. However, 1 year after cessation of transfusion therapy, the direct antiglobu- lin test was negative and the patient's red cells were strongly LW positive. These re-

Allo-Anti LW” 287

sults indicate the patient’s red cells were LW positive and that the LW antigen was only transiently depressed.

Using the proposed terminology ofSiston and Tippett [7] for the LW system, we report a case of normal survival of 7 units of Rho (D)- LW (a+), serologically incompatible red cells in a patient with an allo-anti-LW”. Nor- mal survival had been predicted by MPA and 5’Cr survival studies.

Case History

In August 1979, H.C., a 67-year old Caucasian female was admitted to the hospital suffering from unexplained weight loss and anemia. The patient had no known history of previous transfusions and she had one uneventful pregnancy. No unexpected red cell anti- bodies were detected in the H.C. serum and she was transfused with 1 unit of Rh, (D)’ red cells without incident.

H.C. was again admitted in December 1980 with similar symptoms and right lower-quadrant pain. Work-up revealed a colonic mass with no evidence of metastasis. Carcinoembryonic antigen was 11.1 ng/ml. Her hemoglobin on admission was 6 d d l . Red cell transfusion was requested and at this time an unex- pected red cell antibody, reacting with all red cells of a reagent panel, was detected. Blood samples were refer- red to the Connecticut Red Cross where serologic stud- ies indicated the patient’s serum contained anti-LW”.

H.C. was transferred to the University of Connec- ticut Health Center on December 13 at which time her hemoglobin was 5.2 dd l , hematocrit 17.2% and white blood cell count 21.3 x I O 9 A with a shift to the leR. The patient developed fever and increased abdominal ten- derness requiring surgical intervention for drainage of an abscess. Since LW (a-) blood was not readily avail- able, an in vivo survival of5’Cr labeled, Rho (D)- blood was performed. H.C. received the remainder ofthis unit and three additional Rho (D)-, LW (a+) units without incident. Additional blood was required for a scheduled hemicolectomy. 12 days after the last transfusion, a sec- ond 51Cr survival was performed. Three additional Rh, (D)-, LW (a+) units were successfully transfused.

Materials and Methods

Theantibody in the H.C. serum was identified using a polyspecific antiglobulin reagent (Ortho Diagnostics, Raritan, N.J.) in a standard indirect antiglobulin pro- cedure [2]. Titrations were performed by preparing serial doubling dilution of patient’s serum in saline, incubating at 37°C for 30 min with appropriate reagent red cells, followed by a standard indirect antiglobulin procedure. Reactions were graded and scored [2]. Re- agent red cells were obtained from commercial panels (BCA. Westchester, Pa.) or from liquid nitrogen stor- age.

LW” typings were performed using one example of anti-LW” and one example of anti-LWab in a standard indirect antiglobulin test. All other red cell typings were performed according to manufacturers directions using commercial antisera (BCA, Westchester, Pa. ; Gamma Biologics, Houston, Tex.). Appropriate control red cells were tested in parallel with the H.C. red cells.

The IgG subclass ofthe antibody was determined on V-bottom microtiter plates by an unpublished method [personal commun.; manuscript in prep.]. The method ofSnndkuet al. [4] and Schmfieldet al. [ 5 ] was used for the MPA.

Survival studies were performed infusing 10 ml of donor red cells from Rho (D)- incompatible units la- beled with 50pCi ofsodium chromate (51Cr) by a stan- dard technique [I] . Samples were obtained for the initial study at time 0, 20, 40. 60, 80, 100, 120min and 20h. For the second study, samples were obtained at time 0, 10. 60, 180min and 20h.

Results

Pretransfusion serologic testing yielded the following results. The H.C. red cells were group A, Rho (D)+, rh‘(C)-, rh”(E)+, hr‘(c)+, hr”(e)+. The direct antiglobulin test was negative. The H.C. serum contained anti-LW a, of IgGl subclass, moderately reac- tive (2+) with R,R, red cells and weakly reactive (1+) with rr red cells at the antiglob- ulin phase. Titration of the H.C. serum yielded titers of 128 (score 60) against R ,R , red cells and 32 (score 39) against rr red cells.

288 Cummings/Pisciotto/Roth

Table 1. Results of pretransfusion testing using mononuclear phagocytic assay (MPA)

Patient's serum incubated Percent positive with compared to control

~~

R,R, red cells 20 IT red cells 6

Likelihood of clinically values, % significant extravascular destruction

Low less than 50 High greater than 65 Cannot predict between 5 0 - 6 5

Interpretation: Low likelihood of this example of anti-LWa causing clinically significant extravascular destruction of LW (a+) red cells.

The H.C. red cells did not react with one example of anti-LW". Results of the MPA (table I) indicated a low likelihood of clini- cally significant extravascular destruction of either RIRl or rr red cells.

Rh,, (D)-, LW (a+) red cells were selected for in vivo survival studies since the in vitro reactivity with the patient's serum was weaker than that detected with Rho (D)+, LW (a+) red cells. The initial 51Cr survival study was performed using serologically incom- patible (1 + reactivity at the antiglobulin phase), Rh,, (D)-, LW (a+) red cells that had been collected in CPDA, 4 days previously. Survival was greater than 95% at 1 h and was 78% at 20 h. The patient subsequently re- ceived the remainder of this unit and 3 addi- tional units of Rho (D)-, LW (a+) red cells. All of the transfused units were serologically incompatible yielding a weakly positive (I+) reaction at the antiglobulin phase. Pre- and

posttransfusion hemoglobin and bilirubin levels, as well as antibody titers and reactiv- ity strengths, are shown in figure 1.

14 days after transfusion and following the drainage of the intraperitoneal abscess, hemicolectomy surgery was scheduled and additional blood was requested. Since the strength of the antiglobulin phase reactivity and the titer of the antibody had increased (fig. l), a second 5'Cr survival study of rr erythrocytes was performed. A sample was drawn prior to this study to correct for resid- ual radioactivity from the previous study. Counts taken at 1 and 20 h indicated surviv- als of greater than 95% and 79%, respective- ly. A repeat MPA was not performed at this time.

Prior to surgery, the patient received the remainder of this unit and 2 additional units of Rh, (D)-, LW (a+) red cells that reacted strongly (3') at the antiglobulin phase with her serum. Hemoglobin and bilirubin levels are shown in figure 1.

The H.C. red cells yielded a negative direct antiglobulin test throughout her hos- pitalization. However, circulating LW (a+) red cells could be detected in the posttrans- fusion red cell samples which reacted weakly at the antiglobulin phase with the H.C. se- rum. It therefore appears that in vivo the anti-LW" had a low avidity and easily disso- ciated from LW (a+) red cells.

18 months after transfusion, blood sam- ples from H.C. were retested. Anti-LWa, strongly reactive (3+) undiluted against R,R, and rr reagent red cells, persisted in her serum. Titers and scores decreased to 32 and 45, respectively, against R,R, red cells and 8 and 39, respectively, against IT red cells. The H.C. red cells continued to type LW (a-). The patient gained approximately 30 pounds and felt well with a hemoglobin of

Allo-Anti LWa 289

3

2

2048 1024 512 256 128 6 4 32

.- - 2

u o I

j/ - = - = - -I.- .

abscess drained

.------ ---- > k

:

c

- .- -1: I -- -- I - -

I 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 I 1 1 1 1 1 1 1 1 1 I 1 1 1 1 1 1 1 1 I I

2 / U I5 17 I9 21 23 25 Z? 29 31 2 4 6 8 10 I2 14 16 18 20 22

Fig. 1. The clinical course of patient H.C. who developed allo-anti-LWa is illustrated showing serial hemo- globin, bilirubin and anti-LW a titers following transfusion of Rh, (D)-, LW (a+) serologically incompatible red cells. 0 -0 = R,R, cells; 0 -- 0 = rr cells; numbers in parentheses indicate the strength of reaction at the anti- globulin phase with neat serum.

13 g/dl and a normal carcinoembryonic anti- gen determination.

Discussion

Since this patient required transfusion of large quantities of blood lacking the high incidence antigen LW a, additional tests were performed to determine the efficacy oftrans- fusing red cells that were serologically reac- tive. The use of 5'Cr-labeled red cells as a measurement of red cell survival and deter-

mining in vivo donor compatibility has been well documented [l, 61. Sifvergfeid et al. [6] reported on the predictive value of 5'Cr sur- vival studies using serologically incompati- ble donor red cells in 38 patients. None of the patients were reported to have anti-LW a.

The use of the MPA for determining clinical significance has also been reported by Schanfield et al. [ 5 ] . In that report, the results of the MPA and 5'Cr survival studies avail- able on 8 patients was compared. None of these patients' sera contained anti-LW '.

In the case reported here, both the W r

290 Cummings/Pisciotto/Roth

survival studies and the MPA predicted that this example of anti-LW” had a low likeli- hood of clinically significant extravascular destruction of Rh,, (D)-, LW (a+) red cells.

7 units of Rh, (D)-, LW (a+), serologically incompatible units were subsequently trans- fused without incident. Appropriate incre- ment in hemoglobin levels were seen post- transfusion. The hemoglobin after surgery was decreased due to infusion of fluid during surgery to replace volume loss. The patient was followed for 14 days postsurgery and no additional decrease in hemoglobin was de- tected. The postsurgery total bilirubin in- crease was due to a rise in direct bilirubin, an expected occurrence following this type of surgery. The bilirubin quickly fell to within the normal range.

From all indications, the 7 units of LW (a+) red cells transfused to this patient, did not undergo significant decreased red cell survival, despite the in vitro reactivity of the patient’s serum with the donor red cells. The apparent low avidity of the antibody in vivo, as suggested by the negative direct antiglob- ulin tests and the low percentage macro- phage binding in the MPA, prevented mac- rophage destruction of the transfused red cells.

This is the first reported case of normal survival ofLW (a+) blood in a patient with an allo-anti-LW”. The fact that an increase in the in vitro reactivity (titer and strength of agglutination) of this example of anti-LW a

did not significantly affect in vivo survival, emphasizes the possible lack of correlation between routine serologic tests and biologi- cal activity. When large quantities of rare blood are required because of serologic in- compatibility, the performance of addi- tional tests to determine red cell survival are justified.

Acknowledgements

The authors wish to thank the Special Services Sec- tion of the American National Red Cross for the per- formance of the mononuclear phagocyte assay and the determination ofthe IgG subclass. and Noncy Tower for preparation of the manuscript.

References

I ICSH: Recommended methods for radioisotope red cell survival studies. Br. J. Haemat. 21: 241-250 (1971).

2 Issitt, P. D.; Issitt, C. H.: Applied blood group serol- ogy (Spectra Biologicals, Oxnard IY75).

3 Mollison, P.L.: Blood transfusion in clinical medi- cine (Blackwell, Oxford 1979).

4 Sandler, S.G.; Nusbacher, J.; Schanfield, M.S.: Immunobiology of the erythrocyte (Liss, New York 1980).

5 Schanfield, M.S.; Stevens, J.O.; Bauman, D.: The detection of clinically significant erythrocyte al- loantibodies using a human rnononuclcar phago- cyte assay. Transfusion 21: 571-576 (1981).

6 Silvergleid, A. J.; Wells, R. F.; Hafleigh, E. B.: Korn, G.; Kellner, J. J . : Grumet, F.C.: Compatibil- ity testing using 51 chromium labeled red blood cells in crossmatch positive patients. Transfusion 18:

7 Siston, P.: Tippett, P.: A new allele giving further insight into the LW blood group system. Vox Sang.

8 Stevens, J.O.; Braley, J.F.: Schanfield, M.S.: De- tection ofclinically significant IgG antibodies by an in vitro human peritoneal macrophage phagocyto- sis assay. Transfusion 16: 523 (1976).

9 Tregallas, W. M.; Moulds, J. J.; South, S. F.: Suc- cessful transfusion of a patient with anti-LW and LW positive blood. Transfusion 18: 384 (1978).

8-14 (1978).

42: 252-255 (1982).

Received: January 13, 1983 Accepted: June 29, 1983

E. Cummings, Reference Laboratory, Connecticut Red Cross, Farmington, Conn. (USA)