99 Assessment of Platelet Indices Profile of Pregnant Women Attending University of Abuja Teaching Hospital, Nigeria Amos Dangana1, Anthony Uchenna Emeribe2, Hezekiah Alkali Isah3, Sanusi Musa4, Joel Monday Abu5, Solomon Oloche Onoja6, Nkechi Onukegbe7, Idris Nasir Abdullahi4 1Department of Medical Laboratory Services, University of Abuja Teaching Hospital, PMB 228 Gwagwalada, Abuja Nigeria 2Department of Medical Laboratory Science, Faculty of Allied Medical Sciences, University of Calabar, PMB 1115 Calabar, Nigeria 3Department of Haematology, University of Abuja Teaching Hospital, PMB 228 Gwagwalada, Abuja Nigeria 4Department of Medical Laboratory Science, Faculty of Allied Health Sciences, Ahmadu Bello University, PMB 05 Zaria, Nigeria 5Department of Family Medicine, University of Abuja Teaching Hospital, PMB 228 Gwagwalada, Abuja Nigeria 6Department of Medical Laboratory Science, University of Nigeria, Enugu, Nigeria 7Department of Strategic Information and Research, Institute of Human Virology Correspondence: Idris Nasir Abdullahi, Department of Medical Laboratory Science, Faculty of Allied Health Sciences, Ahmadu Bello University, Zaria, PMB 05 Samaru Road, Zaria, Nigeria Email: eedris888@yahoo.com, inabdullahi@abu.edu.ng Received: May 19, 2021 Revised: July 26, 2021 Accepted: August1, 2019 Published: October 30, 2021 DOI: 10.33086/ijmlst.v3i2.2110 Abstract Platelets initiate hemostasis by aggregating at the site of injury and participate in ensuring endothelial integrity. A defect in this process could lead to intravascular blood loss. This case-control study sought to determine the platelet counts and indices among pregnant women in the University of Abuja Teaching Hospital Gwagwalada, Abuja, Nigeria. A total of 120 pregnant women as case and 60 non-pregnant women as control were enrolled for this study. Blood samples were collected in EDTA tubes, and complete platelet count and indices were carried out using an automated five-part haematology analyzer. The mean ± standard deviation of the platelets count among the pregnant women, 226.54 ± 69.76 109 cells/L was not significantly different from that of the non-pregnant women, 214.95 ± 52.22x 109 cells/L (p=0.295). There was a significant differences in mean platelets volume (MPV) of the case and control groups (p=0.036). After post-hoc test, the significant difference was between the pregnant women in 3rd trimester and the control group (p=0.014). However, there was no diffences in the mena platelets larger cell ratio and platelet distribution width in the case and control groups. Fifteen (11.0%) and 7 (12.1%) of the case and control control groups, respectively had mild thrombocytopenia. However, there was no significant association between pregnant status and thrombocytopenia (p=0.836). Based on these findngs, it can be infered that platelet count and MPV decreases while PDW increase with the progression of gestation age compared to the non-pregnant women. Keywords Platelet indices, Haemostasis, Coagulopathy, Nigeria. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. ©2021 by author. mailto:eedris888@yahoo.com mailto:inabdullahi@abu.edu.ng Ina. J. Med. Lab. Sci. Tech. 2021; 3(2): 99–108 1 0 0 Amos Dangana, et al. INTRODUCTION Pregnancy is a physiological condition and usually does not affect the general health of the pregnant woman. However, pregnancy results in hormonal, hemodynamic and haematological changes (1). These physiological changes need to be considered normal adaptations. Increased total blood volume and hemostatic changes help to combat possible consequences of haemorrhage at delivery (2). It is often argued that altered hemostatic status is required, as the maternal coagulation system prepares for the challenges of parturition, and aims to minimize intrapartum blood loss. However, the alterations in the hemostatic system begin as early as the first trimester, suggesting a requirement for such changes in the proper progression of the early stages of pregnancy, in addition to their role in the regulation of post-partum bleeding (3). For instance, alterations in hemostasis enable the necessary changes in the uteroplacental vasculature to support the establishment of the trophoblast invasion of the spiral arteries of the uterus early in gestations (4). The altered hemostatic status during normal pregnancy presents several physiological challenges in the vasculature and results in an increased risk of excessive thrombosis, especially within the uteroplacental circulation (5). This enhanced pregnancy-associated thrombotic risk may provide the mechanistic basis for many of the major pregnancy complications, such as pre- eclampsia, HELLP syndrome (Hemolysis, Elevated Liver enzymes and Low Platelet count) and intrauterine growth retardation (IUGR) (6). Assessing platelet count in pregnancy is of clinical importance. It is useful in making a diagnosis of HELLP syndrome (7). The HELLP syndrome can often be a complication of pre-eclampsia and eclampsia, determining the severity of these clinical disorders and assessing response level to treatment in HELLP syndrome (8). Pregnancy is associated with endothelial stress and increased platelet aggregation in the uteroplacental circulation resulting in a progressive decline in platelet count with increasing gestational age (9). Furthermore, the increase in plasma volume associated with pregnancy results in dilutional thrombocytopenia (10). Thus, platelet counts are generally lower in pregnancy compared to non-pregnant women, and thrombocytopenia could occur in about 10% of pregnancies (11). Rarely, this may be severe enough to cause maternal and neonatal morbidity and mortality. The elevated platelet aggregation in pregnancy has been attributed to increased formation of thromboxane A2, more intracellular calcium mobilization and reduced synthesis of cyclic adenosine monophosphate (cyclic AMP) (12). With this cognizance, the current prospective study aims to determine the values of platelet Ina. J. Med. Lab. Sci. Tech. 2021; 3(2): 99–108 1 0 1 Amos Dangana, et al. counts and indices among pregnant women attending the University of Abuja Teaching Hospital Gwagwalada Abuja, Nigeria. Data from these could assist to also evaluate pregnant women with potential haemostatic defect and preeclampsia (MPV is useful in this regard). MATERIALS AND METHODS Study Area This study was conducted at the University of Abuja Teaching Hospital (UATH), Gwagwalada, Federal Capital Territory (FCT) Abuja. The laboratory investigations were carried out at the haematology laboratory of UATH. Study Design This was a case-control study on pregnant women who consented to be part of the study at the antenatal clinic at the University of Abuja Teaching Hospital. Controls were enrolled from the female medical students undergoing clinical postings and rotations in the hospital. This study took place from 20th April to 30th December 2018. Selection Criteria for the Case Group Pregnant women who gave written informed consent, and healthy with no history of diabetes mellitus, coagulation- related diseases (such as hemophilia, thalasemia), malaria, HIV/AIDS, Hepatitis-B and C viruses were enrolled as the case subjects. Their biodata alongside parity and gestational ages were extracted from their hospital folders through the assistance of the attending physicians and nurses. Ethics approval Ethical approval was obtained from the ethical committee of the University of Abuja Teaching Hospital (Approval number: UATH / HREC / PR / 2018 / 004.018) Gwagwalada, Abuja, Nigeria. Consent to participate Informed written consent was obtained from all participating subjects before recruiting into the study, following the standards of human experimentation and with the Helsinki Declaration of 1975. Selection Criteria for the Control Group Healthy females (>18 years) who were not pregnant in the last 12 months, with no history of diabetes mellitus, coagulation- related diseases (such as hemophilia and thalasemia), malaria, HIV/AIDS, Hepatitis-B and C viruses were enrolled as the control subjects. They provided their biodata through a short questiionnare administered to them by nurses. Analytical Laboratory Methods Sample collection About 3 millileter (mL) of whole blood samples were collected from all the subjects and dispensed into EDTA (ethylenediamine- N,N,N,N-tetra acetate) tube. Blood samples were analysed in batches within 2 hours of collection until the required number (n=160) was attained. Ina. J. Med. Lab. Sci. Tech. 2021; 3(2): 99–108 1 0 2 Amos Dangana, et al. Platelet Count and Indices The full blood count was carried out using the Genesis HA6000 Automated Hematology Analyzer (calatog number: 9027809900) (Perlong Medical Equipment Company, China). Among other parameters, the analyzer determined platelet count and its various platelet indices (absolute platelet count, mean platelet volume, platelet distribution width and platelet larger cell ratio). Daily and per-run quality control for all the procedures for automated platelet indices analyses were ensured. Statistical Analysis Data were presented as mean and standard deviation on the statistical package for social science (SPSS version 26) and analyzed using student’s t-test and ANOVA for a significant difference in platelet indices between groups. Also, two tailed Chi squared test was used to determine association between 2 catigorical variables. Probability (p<0.05) was used to determine the level of significance for all the statistical analyses. RESULTS In this study, one hundred and eighty (180) participants who met the inclusion criteria were successfully enrolled. Among which 120 were pregnant women (case group) and 60 non-pregnant women (control group). The demographic data obtained for the subjects showed that the most of the control group aged within 21 to 25 years (41.4%) while the majority of the pregnant women (43.2%) were within 26-30 years. Majority of the control group (58.6%) had never been pregnant, whereas (28.0%) of the case group had been pregnant at least once (Table 1). There was a statistically significant difference in age and parity, between the cases and the control groups (p<0.0001) (Table 1). Although, there was an increase in mean platelet count (PLT) in the pregnant women when compared with the non-pregant group (226.54± 69.76 x109 cell/L) versus 214.95±52.22x 109 cell/L), the MPV, PDW, and platelet larger cell ratio were slightly lower in pregnant women than the non- pregnant group. These changes were not statistically different between the 2 study groups (p˃0.05) (Table 2). From our findings, the mean PLT count in the first, second and third trimester of pregnancy (228.64± 45.25 x 109 cell/L, 237.38± 82.39 x 109 cell/L and 212.48± 61.08 x 109 cell/L) did not differ significantly from the mean PLT count of the non- pregannt women (214.95± 52.22 x 109 cell/L) (p=0.184) (Table 3). Moreover, the mean MPV of pregnant women were lowest in 3rd trimester and significantly differ from pregnant women in their 1st, 2nd trimester and the control group (p=0.036) (Table 3). The mean mean platelet volume (MPV) in the first, second and third trimester of pregnancy (10.41± 0.83 fL, 10.33± 0.92fL Ina. J. Med. Lab. Sci. Tech. 2021; 3(2): 99–108 1 0 3 Amos Dangana, et al. and 8.73± 5.70fL) differed significantly from the mean MPV of the non-pregannt women (10.27± 1.81fL) (p=0.036) (Table 3). After post-hoc test, the significant difference was between the pregnant women in 3rd trimester and the control group (p=0.014) (Table 4). However, there was no significant differences in the platelet distribution width and platelet larger cell ratios among pregnant women of all the three trimesters and control groups (p>0.05) (Table 3). Out of the the 180 subjects in this study, 105 (89.0%) and 53 (87.9%) of the case and control groups had normal platelet counts (150-400 x 109 cells/L), respectively. Moreover, 15 (11.0%) and 7 (12.1%) of the case and control groups had mild thrombocytopenia (100-150 x 109 cells/L), respectively. None of the 2 study groups had moderate and severe thrombocytopenia. Also, there was no significant association between pregnancy status and thrombocytopenia (p=0.836) (Table 5). Table 1. Association between age, parity and pregnancy status of study participants Variables Pregnancy Status x 2 p-value Pregnant, n (%) Non-Pregnant, n (%) Age (years) 15-20 0 (0.0%) 13 (19.0%) 52.125 <0.0001* 21-25 16 (11.9%) 24 (41.4%) 26-30 51 (43.2%) 14 (24.1%) 31-35 39 (33.1%) 5 (8.6%) 36-40 14 (11.9%) 4 (6.9%) Parity 0 0 (0.0%) 34 (58.6%) 52.125 <0.0001* 1 33 (28.0%) 14 (24.1%) 2 30 (25.4%) 6 (10.3%) 3 27 (22.9%) 1 (1.7%) ≥4 28 (23.7%) 3 (5.2%) Table 2. Comparison of platelet indices between non-pregnant and pregnant women Platelet Indices Non-pregnant Women (n=60) Pregnant Women (n=120) Mean Difference t value p-value Platelet count (x 109 cells/L) 214.95 ± 52.22 226.54 ± 69.76 -11.59 ±17.54 -1.119 0.295 Mean platelet volume (fL) 10.27 ± 1.81 9.74 ± 3.63 0.53 ±1.82 1.046 0.297 Platelet distribution width 13.60 ± 2.07 13.44 ± 3.60 0.16 ±1.53 0.320 0.749 Platelet larger cell ratio 29.61 ± 7.65 28.38 ± 8.33 1.23 ±0.68 0.972 0.333 Ina. J. Med. Lab. Sci. Tech. 2021; 3(2): 99–108 1 0 4 Amos Dangana, et al. Table 3. Comparison of platelet indices between non-pregnant and the various trimesters of pregnant women Platelet Indices Non-pregnant state (n=60) First trimester (n=24) Second trimester (n=52) Third trimester (n=44) F value p-value Platelet count (x 109 cells/L) 214.95± 52.22 228.64± 45.25 237.38± 82.39 212.48± 61.08 1.632 0.184 Mean platelet volume (fL) 10.27± 1.81 10.41± 0.83 10.33± 0.92 8.73± 5.70 2.913 0.036* Platelet distribution width 13.60± 2.07 13.28± 2.17 13.48± 3.29 13.45± 4.45 0.056 0.983 Platelet larger cell ratio 29.61± 7.65 28.29± 7.74 27.90± 6.89 29.00± 10.12 0.440 0.725 Table 4. Comparison of mean platelet volume between non-pregnant and the various trimesters of pregnant women using the least significant difference (LSD) post hoc Parameter Groups Mean Difference p-value Non-pregnant state First trimester Mean platelet volume (fL) 10.27± 1.81 10.41± 0.83 -0.14± 0.79 0.861 Non-pregnant state Second trimester Mean platelet volume (fL) 10.27± 1.81 10.33± 0.92 -0.06± 0.59 0.917 Non-pregnant state Third trimester Mean platelet volume (fL) 10.27± 1.81 8.73± 5.70 1.55± 0.62 0.014* Table 5. Incidence and severity of thrombocytopenia in non-pregnant and pregnant women Variables Pregnancy Status x2 p-value Pregnant, n (%) Non-Pregnant, n (%) Platelet Count Normal (150-400 x 109 cells/L) 105 (89.0%) 53 (87.9%) 0.043 0.836 Mild (100-<150 x 109 cells/L) 15 (11.0%) 7 (12.1%) Moderate (50-<100 x109cells/L) 0 (0.0%) 0 (0.0%) Severe (<50 x 109 cells/L) 0 (0.0%) 0 (0.0%) DISCUSSION Platelet indices studies in pregnancy have become of great interest owing to the recurrent hypercoagulability crisis accompanied by pregnancy. This study showed that there are changes in the various platelet indices in pregnancy and the mean values of these parameters differ from the control group (non-pregnant women). However, the absolute platelet count (PLT) did not significantly change in pregnant women compared to the contril subjects. This is not in agreement with an earlier study by Boehlen et al. (13) where significant platelet counts decrease was reported among pregnant women. Physiological changes in haematological parameters during pregnancy was reported to have significantly decreased the platelet count in pregnant women (13). Our findings were also not similar to those of Fahmi et al. (14) where they noted that thrombocytopenia was caused by either increased platelet destruction or decreased platelet production. In pregnancy, increased platelet destruction may be mediated by Ina. J. Med. Lab. Sci. Tech. 2021; 3(2): 99–108 1 0 5 Amos Dangana, et al. immunological mechanisms, abnormal platelet activation, or platelet consumption. Increased destruction or utilization of platelets during pregnancy occurs in microangiopathies (affecting small blood vessels) such as thrombotic thrombocytopenic purpura, haemolytic uraemic syndrome, haemolysis, elevated liver enzymes, low platelet (HELLP) syndrome, and pre-eclampsia (14). Another study agrees with the physiologic findings in pregnancy where platelet counts decrease possibly due to haemodilution, which majorly occurs in the third trimester (15). However, the difference in platelet count reported in our study compared to others could be attributed to the nature of sampling techniques or variation in laboratory analytical protocols, as these might have impacted on the results (16). Indeed, there are various hematology analyzers with different technologies and principles of operations (such as the impedance, optical methods and immunofluorescence) current used to count the platelets. Besides, platelet counts are seldom subjected to variations due to artifacts that needed to be ruled out before validation of protocol (16). However, the 5-part automated hematology analyser used in our study is insignifcanly affected by these factors as it uses the latest hematology analysis technology with the best performance caharcteristics (17). These data emphasize the need for additional studies to accurately describe the course of platelet counts throughout normal pregnancies, to document the period platelet counts begin to decrease, and to determine the range of platelet counts among normal women during pregnancy. Findings from this study showed non- significant decreased levels of MPV, PCT and PDW in pregnant women compared to the control subjects (p˃0.05). These changes might be related to the blood volume expansion and hemodilution that occurs during pregnancy. During pregnancy, platelet count decreases gradually from the 1st till the 3rd trimester. In addition, hemodilution has been shown to accelerate platelet consumption which may contribute to a decline in platelet count throughout gestation (9). Interestingly, platelet activation can occur several weeks prior to the clinical onset of pre-eclampsia and increased mean platelet volume (MPV) in the late first trimester of gestation are suggestive of IUGR and PE (9). Our findings was similar to an earlier study by Babah et al. (18), where decreased MPV and PCT in pregnancy was highlighted (18). Findings from our study are also in consonance with that of Nooh and Abdeldayem (19) where they also reported a decrease in the level of MPV, PCT and PDW during pregnancy. Findings from this revealed an increase in the platelet count in the first and second Ina. J. Med. Lab. Sci. Tech. 2021; 3(2): 99–108 1 0 6 Amos Dangana, et al. trimester of pregnancy. However, there was a decrease in platelet count among subject in their 3rd trimester when compared with the control group. These are similar to the findings by Babah et al. (18) where they reported a statistically non-significant increase in platelet count in the first to the third trimester of pregnancy. These observations could be attributed to the feeding habit of the subject (7). It is known that supplements or diet rich in vitamins, iron and folate could enhance erythropoietic activities in the bone marrow, which could assist in boosting platelet counts during pregnancy (20). In most low- and middle- income settings, pregnant women consistent consumption of iron/folate tablets reduces with gestation age (poor adherence) (21, 22). Nevertheless, the progressive fall in platelet count with an increase in gestation did not cause thrombocytopenia in all trimesters of pregnancy as the values were within the normal range (150–450 × 109/L). There was a significant increase in mean platelet volume among subjects in the first and second trimester when compared with the control group. But a significant decrease was observed in the third trimester when compared with the control group. Mean platelet volume (MPV) and platelet distribution width (PDW) were reported to increase during platelet activation (23). Essentially, the decline in the third trimester could be attributed to the physiologic haemodilution. In normal pregnancy, there is often an increase in platelet aggregation and a decrease in the number of circulating platelets with gestation (24). As the platelet lifespan decreases, the MPV increases minimally during pregnancy (24). Thus, the MPV is an accurate measure of platelet size and its considered a biomarker of platelet function (25). Hence, larger platelets with higher MPV counts are reactive and raise higher amounts of the prothrombotic factor thromboxane A2, increasing the tendency to thrombosis (25). The PDW values appeared to have reduced in the three groups of pregnant women, which were statistically non- significant. On the contrary, an increase in PDW was reported by Omorogiuwa and Aigborhuan (26), which was attributed to physiologic compensation for the decreasing platelet count and volume with progression in gestation age (26). Platelets having denser granules are bigger and metabolically active (27), hence PDW is more specific than MPV in the identification of platelet activity. It is a simple and specific marker for enhanced coagulation (23). There was a statistically non-significant increase in the mean of mean± SD of MPV in the first and second trimester among the case group when compared with the mean± SD value of the control group. However, in the Ina. J. Med. Lab. Sci. Tech. 2021; 3(2): 99–108 1 0 7 Amos Dangana, et al. third trimester, there was a significant decrease in MPV among pregnant women. On the contrary, a longitudinal study by Giles (28) showed that MPV increased with gestational age in a statistically non- significant manner. The decline in MPV in the third trimester could be attributed to platelet consumption and hemodilution, conditions common in the third trimester. Findings from this study showed mild thrombocytopenia among the pregnant women when compared with the control group. This explains the normal physiologic changes in pregnancy where hemodilution effects overwhelm the erythropoietic activities of the bone marrow (30). However, pregnant women with thrombocytopenia may be at a higher risk of coagulopathy. CONCLUSION This study is not without limitation. The heterogenous nature of the biodata (especially age) of the case and control groups might signify a bias in subjects selection which could influence the statistics of some analytes. Also, the concurrent analyses of certain platelet factors (such as fibrinogen and Von Willebrand) would have provided more information about the coagulation profile of the pregnant women with low platelet counts and MPV. It is worthy to follow up the pregnant women and neonates with higer risk of developing coagulation complication so that approparite therapeutic interventions could be made available to arrest possible complications. AUTHOR CONTRIBUTIONS Amos Dangana: conceptualization, formal analysis, investigation, supervision, writing - original draft, writing - review & editing. Anthony Uchenna Emeribe: formal analysis, methodology, writing - original draft, writing - review & editing. Hezekiah Alkali Isah: writing - original draft, writing - review & editing. sanusi musa: writing - original draft, writing - review & editing. Joel Monday Abu: writing - original draft, writing - review & editing. Solomon Oloche Onoja: writing - original draft, writing - review & editing. Nkechi Onukegbe: writing - original draft, writing - review & editing. Idris Nasir Abdullahi: conceptualization, formal analysis, investigation, methodology, supervision, writing - original draft, writing - review & editing. CONFLICT OF INTEREST None declared by authors. REFERENCES 1. Capasso G, Unwin R. Electrolytes and acid–base: common fluid and electrolyte disorders. Medicine (Baltimore) [Internet]. 2011 Jun;39(6):317–24. Available from: https://linkinghub.elsevier.com/retrieve/pii/S135 7303911000636 Ina. J. Med. Lab. Sci. Tech. 2021; 3(2): 99–108 1 0 8 Amos Dangana, et al. 2. Rahmawati F. Aspect of chronic kidney disease. J Ilm Kedokt Wijaya Kusuma. 2018; 3. Makani M, Setyaningrum N. Patterns of furosemide use and electrolyte imbalance in heart failure patients at Hospital X Yogyakarta. J Ilm Farm. 2017; 4. Tyas RA, Damayanti W, Arguni E. Prevalence of serum electrolyte disorder in children under five with diarrhea and dehydration in dr sardjito hospital on 2013-2016. Sari Pediatr. 2018; 5. Kusnanto. Modul learning modules meeting fluid and electrolyte needs. Fakultas Keperawatan Universitas Airlangga. 2016. 6. Yaswir R, Ferawati I. Physiology and balance disorders of sodium, potassium and chloride and laboratory examination. J Kesehat Andalas. 2012; 7. Horne M dan PS. Pocket guide to fluids, electrolytes, and acid-base balance. 2nd ed. Asih Y, editor. Jakarta: EGC; 2012. 8. Pfortmueller CA, Uehlinger D, von Haehling S, Schefold JC. Serum chloride levels in critical illness—the hidden story. Intensive Care Med Exp. 2018; 9. Salam SH. The basics of fluid and electrolyte therapy. Bahan Kuliah FK Unhas. 2016; 10. Plebani M. Quality indicators to detect pre- analytical errors in laboratory testing. Clinical Biochemist Reviews. 2012. 11. Lippi G, Chance JJ, Church S, Dazzi P, Fontana R, Giavarina D, Grankvist K, Huisman W, Kouri T, Palicka V, Plebani M, Poro V, Salvagno GL, Sndberg S, Sikaris K, Watson I, Stankovic AK, Simundic AM. Preanalytical quality improvement: From dream to reality. Clin Chem Lab Med. 2011 Jul;49(7):1113-26. 12. Baruah A, Goyal P, Sinha A, Ramesh KL, Datta R. Delay in specimen processing-major source of preanalytical variation in serum electrolytes. J Clin Diagnostic Res. 2014; 13. Trisna YN, Mutmainnah M, DN Pakasi R, Hardjoeno H. Na, K, Cl concentration in time interval examination variations of serum. Indones J Clin Pathol Med Lab. 2018; 14. Balci AK, Koksal O, Kose A, Armagan E, Ozdemir F, Inal T, Oner N. General characteristics of patients with electrolyte imbalance admitted to emergency department. World Journal of Emergency Medicine. 2013;4(2):113-6 15. Arneson W. Electrolytes: The salts of the earth. Laboratory Medicine. 2014. 16. Barth JH. Clinical quality indicators in laboratory medicine. Ann Clin Biochem. 2012; 17. Roumelioti M-E, Glew RH, Khitan ZJ, Rondon- Berrios H, Argyropoulos CP, Malhotra D, et al. Fluid balance concepts in medicine: Principles and practice. World J Nephrol. 2018; 18. Hedayati M, Razavi SA, Boroomand S, Kheradmand Kia S. The impact of pre-analytical variations on biochemical analytes stability: A systematic review. Journal of Clinical Laboratory Analysis. 2020. 19. Donnelly JG, Soldin SJ, Nealon DA, Hicks JM. Stability of twenty-five analytes in human serum at 22 degrees C, 4 degrees C, and -20 degrees C. Pediatr Pathol Lab Med. 1995; 20. O’Keane MP, Cunningham SK. Evaluation of three different specimen types (serum, plasma lithium heparin and serum gel separator) for analysis of certain analytes: clinical significance of differences in results and efficiency in use. Clin Chem Lab Med [Internet]. 2006 Jan 1;44(5). Available from: https://www.degruyter.com/view/j/cclm.2006.44. issue-5/cclm.2006.099/cclm.2006.099.xml 21. Dupuy AM, Cristol JP, Vincent B, Bargnoux AS, Mendes M, Philibert P, et al. Stability of routine biochemical analytes in whole blood and plasma/serum: focus on potassium stability from lithium heparin. Clin Chem Lab Med [Internet]. 2018 Feb 23;56(3):413–21. Available from: https://www.degruyter.com/doi/10.1515/cclm- 2017-0292 22. Scott MG, LeGrys VA, Hood JL. Electrolytes and blood gases. In: Tietz Textbook of Clinical Chemistry and Molecular Diagnostics. 2012. 23. Burger P, Kostova E, Bloem E, Hilarius-Stokman P, Meijer AB, van den Berg TK, et al. Potassium leakage primes stored erythrocytes for phosphatidylserine exposure and shedding of pro- coagulant vesicles. Br J Haematol. 2013; 24. Apriliani I, Santosa B, Sukeksi A. Difference in electrolyte levels (Na, K, Cl) in samples immediately and delayed by 150 minutes. repository.unimus.ac.id. 2018. 25. Azizah N, Aliviameita A. Effect of delay of serum examination on electrolyte levels of sodium and chloride. J Med Lab Sci Technol. 2019; 26. An B, Park C-E. Evaluation of stability of serum on different storage temperatures for routine chemistry analytes. Korean J Clin Lab Sci. 2014; 27. Departemen Kesehatan RI. Pedoman praktek laboratorium yang benar (Good laboratory practice). Jakarta; 2008.