Pseudotrombocytopenia Caused by EDTA in Obstetrics and Perinatology – Case Report A

1 Hematology and Blood Transfusion Laboratory, Gynecology and Obstetrics Clinic, Clinical Center, Belgrade, Serbia 2 Department of Pregnancy Pathology, Clinic of Gynecology and Obstetrics, Clinical Center, Belgrade, Serbia 3 Department of children born with Cesarean section, Clinic for Gynecology and Obstetrics, Clinical Center, Belgrade, Serbia 4 Department of Anesthesiology and Reanimatology, Clinic of Gynecology and Obstetrics, Clinical Center, Belgrade, Serbia


INTRODUCTION
Pseudothrombocytopenia is a phenomenon in which antibodies, present in the patients' blood react with platelets in blood sample anticoagulated with EDTA, causing agglutination and a spuriously low platelet count [1].Although EDTA-dependent pseudothrombocytopenia has been practically and literally well-handled for more than 30 years, today it may also be unrecognizable.
Pseudothrombocytopenia most commonly occurs when using anticoagulant Ethylene Diamine Tetra Acetic acid (EDTA) in blood laboratory sampling.Clinically, the most important, pseudothrombocytopenia occurs in pregnant women and in their newborns if blood sample is taken using EDTA [2].Analyzation of platelets could be done from the newborn sample taken with another anticoagulant (ammonium oxalate, citrate) and tested immediately aft er sampling [2].Th is condition in a newborn disappears aft er a month of birth [3,4,5].

CASE REPORT
Patient MT, 35 years old, obese pregnant woman 36 weeks of gestation who got pregnant naturally for the second time, was admitted to Gynecological and Obstetrics Clinic Clinical Centre of Serbia for monitoring pregnancy and delivery planning.
Previous natural pregnancy was due to the pelvic presentation of the fetus delivered by Cesarean section.Th e fi rst child was born healthy.
During this second pregnancy was administered antihypertensive (methyl-dopa) and anticoagulant therapy (sirete fraxiparine) due to the comorbid clinical conditions hypertension and varices cruris et femoris bilateralis.In addition, laboratory values of platelets were monitored.
Due to obstetric reasons such as:pelvic presentation of the fetus, intrauterine growth restriction (IUGR) and reduced amounts of amniotic fl uid (oligoamnion) has been determined to deliver a child with the Cesarean section at 36 th week of gestation.Th e cardiotocography (CTG) record was within normal limits.
EDTA pseudothromocytopenia was suspected considering that at the admission, the EDTA platelets (Plt) result was 34.0 x10e9/L (MPV 16, 9fl ) and patient was without any symptoms of bleeding tendency.We decided to check it by examining blood smear and incubating EDTA sample on 37°C for 1 hour.Th e Plt value on 37°C was 32.0 x 10e9/L and blood smear showed a platelet aggregation (Figure 1).
Blood samples were simultaneously taken in four tubes containing four diff erent types of anticoagulants; EDTA, Sodium Citrate (Na-citrate), Amonium oxalate and Magnesium-sulfate (Mg-sulfate).Th e blood samples were run in fully automated Hematology analyser Siemens ADVIA 2120i.Th e platelet count www.hophonline.orgAdditionally, blood sample on EDTA was measured every 10 minutes from the moment of taking, and results were: 156.0 x Th e decision was made to terminate the pregnancy with the Cesarean section so the patient was on prophylactic therapy with Low molecular weight heparin (LMWH) (Nadroparin-calcium, Fraxiparine®, 2850 i.j./ 0.3 ml sc).Because of the possibility of developing heparin-induced thrombocytopenia (HIT), antibodies to the heparin / PF4 complex were controlled and result was negative.HIT antibodies were determined by using Pa-GIA Heparin / PF4 Antibody test kit, BioRad Switzerland.
Despite the results of the laboratory and the precise transfusiologists' diagnosis of the pseudothrombocytopenia, the surgical team requires preparing 2 pools of platelet concentrates.Due to the careful monitoring of transfusiologists, platelet concentrates were not administered to the patient.
Newborn was healthy boy (Apgar score 9, length 48 cm, body weight 2700 g, head circumference 35 cm. Th e newborn's Complete Blood Count (CBC) results revealed normal parameters apart from a low platelet count 49.0 x 10e9/L (MPV 9,4 fl ).He had no bleeding tendency.A blood smear was ordered and it revealed platelet aggregations (Figure 4.).
EDTA.Literate data indicate the possibility of pseudothrombocytopenia in children born by mothers with pseudothrombocytopenia, as in our case.It is recommended that blood smear sample should be mandatory for that children.

DISCUSSION
Gowland and his team fi rst reported in 1969 about PTP induced by EDTA (6).
EDTA induced alteration of surface glycoproteins (GP) enables binding of antiplatelet antibodies.Th is is the cause of agglutination.Platelet agglutinins could belong to all immunoglobulin classes (IgG, IgM, IgA) (7,8).Among these 20% of EDTA induced PTP show the agglutination in citrate anticoagulant as well.Oxalate and heparin have also been implicated in PTP (7,9).
Th is phenomenon in a newborn disappears aft er a month of birth indicating transplacental transmission of plasma components (most likely IgG) leading to pseudothrombocytopenia of the newborn (10).Due to this phenomenon, concerning neonate with asymptomatic thrombocytopenia shall be proceeded with patterns as adults in order to avoid inappropriate and potentially harmful treatment decisions [3,4,5,11].
In addition to enable better diagnostic and therapeutic approach needed for personalized medicine concept, it is necessary to include cpecialist of clinical transfusiology, clinical pharmacology and clinical immunology in every day practice.To aquire this level it is necessary to enlarge these specialists number.

CONCLUSION
Although PTP does not require clinical-therapeutic intervention, undiagnosed PTP can lead to therapeutic intervention (unnecessary administration of platelet concentrate) and seriously deteriorate both mother's and newborn's health.Th erefore, the authors suggest examination of blood smear in every case when thrombocytopenia is detected and warn to respect multidisciplinary medical team work.Newborn platelet counts from a blood sample taken from sodium citrate was 189.0 x 10e9 / L (MPV 7.3 fl ), which made it clear that it was pseudothrombocytopenia.So, the baby was considered to have pseudothrombocytopenia and discharged from the hospital.
It has been advised that the baby's platelets should be controlled one month after birth due to the apparent elimination of the transplacental cause of pseudothrombocytopenia.Finally, during his last follow up visit, he was 2 months old and the baby's platelet count was normal, even in the presence of

Figure 1 .Figure 2 .
Figure 1.Peripheral smear sample of mother's blood taken on EDTA, A x 400 B x 1000

Figure 3 .
Figure 3. Mother's platelet values measured in a blood sample on EDTA every 10 minutes from the moment of blood sampling

Figure 4 .
Figure 4. Blood smear of the newborn's peripheral blood sample taken on EDTA