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Identifying False Positive Weak Rh (D) agglutination test in DAT positive red cells

Identifying False Positive Weak Rh (D) agglutination test in DAT positive red cells
#00063175
Author: Reshma Nambiyar, MBBS; Manish Raturi, MBBS, MD; Anuradha Kusum, MBBS, MD
Category: Laboratory Hematology
Published Date: 08/24/2020

Short description:

a) A 55-year-old male diagnosed with acute liver disease [? Autoimmune pathology] was admitted to the emergency room of our hospital for yellowish discoloration of his urine and eyes since last three days. He also complained of generalized weakness and loss of appetite for the last four days. There was accompanying breathlessness on exertion. There was no history of fever, hypertension and or diabetes mellitus.

b) On investigating, his hemoglobin and platelet counts were 4.6g/dL and 95000 per mm3 respectively. His low blood counts warranted an urgent transfusion of packed red blood cells.

c) His EDTA sample showed a blood grouping discrepancy by the conventional tube technique [CTT]. While, his cell type showed B Rh (D) mixed field (Mf) weak reaction, his serum showed varying grades of agglutination with in-house prepared A, B and O pooled cells interpreted as 4+, 1+ and 1+ respectively. His direct agglutination test [DAT] and autologous control both were positive. On keeping the test tubes of his serum grouping at 37°C for one hour, the 1+ grade reaction in B and O pooled cells were resolved. His ABO status was, therefore, confirmed to be B type. His Rh (D) status was retested using Anti-Rh (D) antisera of two different manufacturers and macroscopically both showed a weak reaction in the CTT.

d) We checked for his anti-Rh (D) agglutination microscopically and confirmed it to be an Mf reaction with Anti-D [IgM] under 40x magnification [fig (a)]. Additionally, his weak D testing on using Anti-D (1:32 diluted IgG) followed by the addition of anti-human globulin reagent showed false positive [FP] agglutinates [fig (b)]. Therefore, accurate determination of his D variant status could not be done serologically due to his DAT positive red cells [that gave FP agglutination, leading to invalid results].

e) His blood typing was eventually interpreted as B Rh (D) [Mf? D variant] with the following transfusion advice;

·         For red blood cell transfusion kindly consider B or O Rh (D) negative donor cells

·         If fresh frozen plasma and or platelets are required, either B or AB plasma may be considered.

f) The weak Rh (D) phenotype is a weakened expression of the Rh (D) antigen due to fewer Rh (D) antigens per red cell than normal Rh (D) positive red cells [1]. It is, therefore, prudent to consider individuals with a weak Rh (D) antigen as Rh (D) positive when presenting as a donor and Rh (D) negative when confronted as a recipient [2].

g) Patient was given two units of best matched O Rh (D) negative packed red cell transfusions apart from supportive treatment. The patient’s relatives were unwilling to continue the treatment and got him discharged against medical advice. We eventually lost the patient to any further follow-up.

References:

1. The Weak D (Du) Phenotype. Available from: https://sites.ualberta.ca/~pletendr/tm-modules/rh/70rh-weakd.html   [last accessed on 20.08.2020]

2. Kumar H, Mishra DK, Sarkar RS, Jaiprakash M. Difficulties in Immunohaematology: The Weak D Antigen. Med J Armed Forces India. 2005 Oct;61(4):348-50.