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Year : 2022  |  Volume : 13  |  Issue : 1  |  Page : 35-40

The prevalence of cumulative alloimmunization in patients with sickle cell disease at King Fahad University Hospital

Department of Pathology, Imam AbdulRahman Bin Faisal University, IAU University, King Fahad University Hospital, Dammam; Laboratory and Blood Bank, Blood Bank, Ministry of Health, Qatif Central Hospital, Qatif, Saudi Arabia

Date of Submission24-Aug-2021
Date of Acceptance13-Dec-2021
Date of Web Publication28-Apr-2022

Correspondence Address:
Dr. Rabab Ahmad AlDawood
P.O Box: 4332, Dammam, 34243
Saudi Arabia
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/joah.joah_122_21

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BACKGROUND: Sickle cell disease (SCD) is caused by a mutation in the beta-globin gene. Red blood cell (RBC) transfusion is considered the mainstay of management. On the other hand, it carries many side effects, of which alloimmunization is the most significant.
AIMS AND OBJECTIVES: The aim of this study is to identify the prevalence of alloimmunization among SCD patients and its relation with other independent risk factors in order to provide recommendations for the care of SCD patients.
MATERIALS AND METHODS: This is a retrospective cohort study conducted at King Fahad Hospital of the University in Al-Khobar from January 1, 2010, to December 31, 2018. Data were collected from both the QuadraMed electronic system and the blood bank transfusion cards. Data were analyzed using IBM SPSS (version 23.0, Chicago, IL, USA).
RESULTS: One hundred and seven out of 556 SCD patients developed alloantibodies with a prevalence of (19.2%). Anti-E was the most identified alloantibody in 37 (34.6%) patients, followed by anti-K in 33 (30.8%) of the alloimmunized SCD patients. We found a clinically significant correlation between the alloimmunization and age and direct antiglobulin test positivity.
CONCLUSION: Alloimmunization is a major complication among SCD patients. Moreover, most of the formed alloantibodies were directed toward the Rh and K antigens. All SCD patients should undergo extended RBC phenotyping at the earliest opportunity, including the following RBC antigens (C/c, E/e, K, Jka/Jkb, Fya/Fyb, M/N, and S/s) at a minimum either serologically or by genotyping.

Keywords: Alloimmunization, blood transfusion, sickle cell disease

How to cite this article:
AlDawood RA. The prevalence of cumulative alloimmunization in patients with sickle cell disease at King Fahad University Hospital. J Appl Hematol 2022;13:35-40

How to cite this URL:
AlDawood RA. The prevalence of cumulative alloimmunization in patients with sickle cell disease at King Fahad University Hospital. J Appl Hematol [serial online] 2022 [cited 2022 Jun 25];13:35-40. Available from: https://www.jahjournal.org/text.asp?2022/13/1/35/344254

  Introduction Top

Hemoglobinopathies are diseases involving the hemoglobin (Hb) molecule, all hemoglobinopathies result from genetic mutations affecting the Hb molecule either quantitatively or qualitatively.[1] Sickle cell disease (SCD) is the term used to describe the homozygous inheritance of two HbS molecules (sickle cell anemia) or the heterozygous combination of one HbS together with another variant affecting the beta-globin chain as HbC or beta-thalassemia.[1] HbS is caused by a point mutation in nucleotide 17 replacing the original adenine with thymine, resulting in the substitution of valine for glutamic acid at amino acid position six in the beta chain, the inheritance pattern is autosomal codominant.[1] This substitution will change the size and charge of the original amino acid; therefore, folding of the affected chain and the interactions with the adjacent amino acids will be altered.[1] The consequences of this monogenic mutation are acute and chronic complications with multisystem involvement such as vaso-occlusive sickle crises, infections, multisystem organ failure, and chronic organ damage.[1] The prevalence of SCD in Saudi Arabia was estimated to be 4.5% from the premarital screening program published in 2018.[2] Furthermore, the Eastern Province was the most prevalent, with 9.8 cases per 1000 population.[2]

Although red blood cell (RBC) transfusion is an important modality of treatment for many acute and chronic complications, it predisposes to many side effects such as iron overload and RBC alloimmunization.[3] Alloimmunization can be defined as an immune response against foreign RBC antigens from blood transfusions. The prevalence of RBC alloimmunization in SCD patients in Saudi Arabia is estimated to be 23.9%.[4]

In general, the antigenic differences between the donor and the recipient are the initial trigger of forming an alloantibody.[5] Other risk factors associated with higher alloimmunization among SCD are the higher cumulative number of units received, older age, age at first lifetime transfusion, female gender, inflammatory state, and variant Rh alleles.[6] Conversely, not all transfused SCD patients will form alloantibodies, and this can be explained by variability of genetic susceptibility toward alloimmunization resulting in the increased tendency to form multiple alloantibodies known as “responder” phenotype.[7]

The development of alloantibodies will add to the complexity of finding a compatible blood unit and may result in delayed hemolytic transfusion reactions (DHTRs). This is classically defined as an anamnestic reaction caused by a previously existing alloantibody performed against specific RBC antigens which was undetectable at the time of transfusion. The features of DHTR are variable from mild to severe and can be life-threatening predisposing to hyperhemolytic crisis defined as hemolysis of both transfused and autologous RBCs.[8]

The standard of care in RBC transfusions nowadays is with partial extended matching for Rh (D, C, E, c, e) and K for many centers around the world.[9] However, until the end of 2013, King Fahad Hospital of the University (KFHU) in Al Khobar was found to follow a strategy where the transfusion is matching only for ABO and D antigens. Moreover, a retrospective study was published in 2007 and conducted at KFHU regarding the prevalence of alloimmunization in SCD patients. The study found a frequency of 13.7%.[10]

The aim of this study is to determine the prevalence of alloimmunization among SCD patients at KFHU from 2010 to 2018 and correlate alloimmunization risk with other independent risk factors as cumulative number of transfused units, gender, age, age at first lifetime transfusion, SCD phenotype, and direct antiglobulin test (DAT).

  Subjects and Methods Top

Study design and population

This is a retrospective cohort study conducted at KFHU in Al-Khobar. The study includes all SCD patients from January 1, 2010, to December 31, 2018. Excluding SCD patients who did not receive a blood transfusion or those in whom their transfusion history could not be traced at our institution.

Data collection

Upon the approval of Institutional Review Board (IRP-2019-01-352), data were collected from medical records (QudraMed electronic system) and transfusion cards from the blood bank. Hb electrophoresis is used to identify the SCD phenotypes, and from the transfusion cards, the cumulative number of transfusions, age of first transfusion, and type of alloantibody were obtained.

Laboratory investigations

After ABO, Rh, and K blood grouping by ID-Card DiaClon ABO/D + reverse grouping (DiaMed, Cressier, Switzerland) and ID-Card DiaClon Rh subgroups + K (DiaMed, Cressier, Switzerland), respectively, the following were done routinely for every patient.

Antibody screening

Direct anti-human globulin (AHG) test was done using ID-Card Low-Ionic Strength Solution. The patient serum/plasma was incubated with three test cell reagents (I-II-III) at body temperature. Providing a positive AHG, further testing for antibody identification will be performed.

Antibody identification

This is performed to identify unexpected antibodies detected in the antibody screening, using ID-gel card method with 12 commercial cell panels of adult RBCs against the patient's serum.


A cross-match was performed after the selection of the appropriate blood from the blood group results. First, a saline procedure was performed at room temperature, then at 37°C, followed by an antiglobulin phase using albumin or low-ionic strength solution. Only the units with compatible cross-matching (i.e., antigen negative) were chosen for transfusion.

Direct antiglobulin test

DAT is performed using the same ID-Card low-ionic strength solution used in antibody screening without the incubation step to detect the presence of any antibody or complement coating the RBCs through polyspecific immunoglobulin G (IgG). Positive results will go for further testing using monospecific IgG AHG reagents (ID-Card Coombs Anti IgG).

Data analysis

The data were analyzed using IBM SPSS (version 23.0, Chicago, IL, USA). Descriptive statistics such as frequency and percentages or median and interquartile range were used according to the variable type. For all the categorical variables, Chi-square or Fisher's exact test were used to compare between the alloimmunized and nonalloimmunized patients and Mann–Whitney test was used for all continuous variables. Logistic regression analysis was used to determine the risk associated with alloimmunization. P < 0.05 was considered statistically significant.

  Results Top

A total of 556 SCD patients who received at least one transfusion were included in the study, of which 107 patients developed alloantibodies with a prevalence of 19.2% (95% confidence interval [CI] [16.0–22.8]). Of the 107 alloimmunized patients, there were 46 (8.3%) patients having a positive antibody screening from the first encounter, possibly from a previous RBC transfusion outside our institution. There were 274 (49.3%) males and 282 (50.7%) females. Patients were mainly Saudis 535 (96.2%) with homozygous HbSS 469 (84.4%). The median age of the patients was 29.8 years (interquartile range [IQR]: 20.9–36.3).

For the demographic and transfusion characteristics for all SCD patients, the most common blood group in order was O (n = 287, 51.6%), A (n = 143, 25.7%), B (n = 106, 19.1%), and AB (n = 20, 3.6%). Rh positive was most common in 521 (93.7%) patients. The DAT was positive in 60 (10.8%) patients and was negative in 496 (89.2%) patients. The median age at first transfusion of the patients was 21.0 years (IQR: 9.5–28.3). The median cumulative number of units received by the patients was 4.0 units (IQR: 2.0–12.0) [Table 1].
Table 1: Demographics and transfusion data for all sickle cell disease patients in the study

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Anti-E was the most identified alloantibody in 37 (34.6%) patients, followed by anti-K in 33 (30.8%) of alloimmunized SCD patients [Table 2]. Among the 107 alloimmunized patients, 71 (66.4%) were identified with one antibody, 21 (19.6%) were identified to have two antibodies, ten (9.3%) with three antibodies, and five (4.7%) were identified with more than three antibodies.
Table 2: Prevalence and specifications of the alloantibodies identified among sickle cell disease patients

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The variables such as gender, nationality, blood group, Rh, and SCD phenotype did not show a significant association with alloimmunization. The alloimmunized patients showed a higher proportion of positive DAT compared to nonalloimmunized patients (63.3% vs. 13.9%, P = 0.000). In addition, the age and the age at first unit transfusion of the alloimmunized patients were significantly higher than the nonalloimmunized patients (P = 0.000, P = 0.037), respectively. Furthermore, the cumulative number of units received by the alloimmunized patients was significantly more than the nonalloimmunized patients (P = 0.000).

The logistic regression analysis shows that increasing age was associated with an increased likelihood for developing alloimmunization (odds ratio [OR] = 1.08, 95% CI [1.0, 1.1]). The odds for a patient to develop alloimmunization was more likely in patients with a positive DAT as compared to those with a negative DAT test (OR = 10.23, 95% CI [5.5, 18.9]). Increasing age at first transfusion was associated with a reduction in the odds of developing alloimmunization (OR = 0.954, 95% CI [0.9, 1.0]) [Table 3].
Table 3: Logistic regression analysis of alloimmunization and its correlates

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  Discussion Top

Blood transfusion therapy is the cornerstone in SCD management, and the development of alloantibodies will further complicate the disease. For that reason, this study aimed to highlight the prevalence of alloimmunization among SCD patients. In the current study, 107 out of 556 patients have developed alloantibodies with a prevalence of 19.2%. This prevalence was higher than a previously published study in our institution in 2007 which revealed a risk of 13.7%,[10] this could be explained by the possible increased use of RBC transfusions both episodic and chronic to reduce the disease morbidity and mortality, together with the fact that most of our patients in the study have received blood transfusion prior to the implementation of partially extended blood phenotyping. However, the mentioned prevalence in this study was comparable to other studies published in Saudi Arabia,[11] and within the range of more international published studies.[6],[12],[13],[14]

Furthermore, antibodies to RhE, K, RhC, and Rhc were the most encountered, with a prevalence of 34.6%, 30.8%, 14%, and 11.2% of alloimmunized SCD patients, respectively. This result matched most of the studies in the literature,[6],[10],[11],[15],[16],[17],[18] in agreement with the result of Evers et al. about their immunogenicity and potential to stimulate the humoral immune response.[19] This provides further evidence that strict prophylactic partial RBCs matching for Rh and K antigens for all SCD patients might reduce the incidence of alloimmunization,[20] unless this alloimmunization was related to Rh variability undetected by serological testing method. On the other hand, other alloantibodies to other blood group systems are still encountered, mainly the Duffy, Kidd, and MNS systems with a prevalence of alloimmunization ranging from 1.9% to 6.5%, and matching for those RBC antigens may possibly further reduce the alloimmunization risk.

The cumulative number of units transfused was significantly higher in alloimmunized SCD patients compared to nonalloimmunized in the bivariate analysis with a median of ten and four RBC units, respectively, in consistence with other published studies.[6],[12],[14],[17],[19],[21] However, the odds in the multivariate analysis did not show any correlation. On the other hand, alloimmunized SCD patients demonstrated the tendency for developing multiple antibodies, 21 (19.6%) patients were identified to have two antibodies, ten (9.3%) patients with three antibodies, and five (4.7%) patients were identified with more than three antibodies, in consistence with the same finding in other studies,[6] emphasizing that host-related risk factors play a role. Another independent risk factor associated with alloimmunization is the patient's age with significantly higher alloimmunization risk in the older age group. This finding was previously highlighted by similar articles[11],[17] which showed the same finding. On the contrary, although alloimmunized SCD patients had a significantly older age at their first transfusion, the odds logistic regression revealed no difference, unlike some of the published studies in the literature,[6] this could be explained by the fact that most of our SCD patients have been transfused elsewhere at a younger age. However, some reports concluded with a similar result that showed no relation between the age of first transfusion and alloimmunization.[18] Furthermore, females had a higher prevalence of alloimmunization than males, in agreement with some reports[10],[11],[14],[15],[22] and contrary to others.[17] Nevertheless, considering female gender as an independent risk factor for alloimmunization is arguable due to pregnancy and delivery exposure adding to their alloimmunization risk. Interestingly, positive DAT was estimated to be (8.3%) for all SCD patients and (63.3%) of the alloimmunized, the same finding has been addressed by Aygun et al. with (17.7%) positive DAT for both adult and pediatric SCD populations in addition to that 75% of them had a prior alloantibody.[13] This is a well-known phenomenon where DAT can be positive with an unclear explanation but will further add to the complexity of pretransfusion testing.[23] In a review article published in 2017 by Parker and Tormey. it was mentioned that a positive DAT result in a patient with a previous transfusion history could be the first indication of developing an alloantibody;[24] furthermore, DAT can be used to detect the antibodies targeting the transfused RBCs before hemolysis is evident or the presence of an alloantibody in a suspected case of delayed hemolytic transfusion reaction.[24] This fact does not necessitate the presence of an autoantibody, an elution is necessary to demonstrate the possible alloantibody presented in a low titer in the plasma and not yet detectable or the presence of new pan-reacting autoantibody; nonetheless, a positive DAT may occur without any clinical manifestation or hemolysis.[25] Finally, our result did not show any correlation between alloimmunization risk and other factors as nationality, blood group, Rh, and SCD phenotype, this was consistent with another study.[17]

This study has some limitations, as most of our patients have an untraceable transfusion history outside our institution with the possibility of different transfusion protocols over time; hence, our comparison with other studies is limited as we do not have a national transfusion database. A second limitation is that some patients have received blood transfusions at our institution with no further antibody screening posttransfusion, this might underestimate the alloimmunization risk. Third, as the study included both electronic and paper-based records; those with missing records were excluded. Furthermore, we did not have the RBC unit phenotype before transfusion to conclude possible Rh variability in cases developing alloantibodies despite phenotypic matching.

  Conclusion Top

RBC alloimmunization is a major complication in SCD management, mainly toward Rh and Kell blood group systems with the tendency of developing multiple alloantibodies. We found a clinically significant correlation between alloimmunization and age and DAT positivity.

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Conflicts of interest

There are no conflicts of interest.

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  [Table 1], [Table 2], [Table 3]


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