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Uvod: Krioglobulini su cirkulirajući imunoglobu- lini ili imunoglobulinski kompleksi koje karakterizira reverzibilna, hladnoćom izazvana precipitacija. Krioglobulini uzrokuju analitičku interferenciju prilikom mjerenja parametara krvne slike na hematološkim analizatorima (HA), što dovodi do lažno povećanih rezultata, uglavnom broja Lkc i/ili Trc. Prikaz slučaja: 82-godišnji muškarac primljen je u našu bolnicu zbog purpure, slabosti, oticanja u obje noge i značajne proteinurije i hematurije. Tijekom boravka u bolnici, pacijentu su kvalitativno detektirani krioprecipitati u serumu te su uočene neobjašnjive promjene u broju Trc tijekom nekoliko uzastopnih mjerenja. Rezultati: Prikazujemo rezultate istog uzorka izmjerenog na Advia 2120i HA u istom danu, prema sljedećem redoslijedu: 1) odmah po prijemu u laboratorij: TRC 158 x10 9/L, MPV 10, 0 fL ; 2) 1, 5h po prijemu, čuvano na sobnoj temperaturi (ST): TRC 314 x109/L, MPV 14 fL ; 3) nakon zagrijavanja na 37 °C 30 min i neposrednog mjerenja: TRC 132 x10 9/L, MPV 9, 1 fL ; 4) 45 min nakon zagrijavanja, čuvano na ST:TRC 279 x109/L, MPV 13, 6 fL ; 5) 3h nakon zagrijavanja, čuvano na ST: TRC 398 x109/L, MPV 16, 3 fL. Trc citogram je bio prepun čestica u području šuma trombocitnog porijekla i HA je generirao pripadajuće upozorenje. Histogram distribucije trombocita po volumenu je pokazivao rastezanje prema većem volumenu kod uzoraka koji su stajali na ST. Razmazi venske krvi napravljeni su neposredno nakon mjerenja 3, 4 i 5 te obojani MGG tehnikom. Uočene su ekstracelularne nakupine blago ružičastog amorfnog materijala u razmazima 4 i 5. Zaključak: Prepoznavanje interferencija povezanih s krioglobulinima kod mjerenja parametara krvne slike može biti vrlo izazovno u rutinskoj laboratorijskoj praksi. Njihova interferencija kod HA ovisi o veličini, nije dosljedna ili relativna u odnosu na količinu krioglobulina ili njegovu prirodu. Svako odstupanje u broju Trc ili Lkc u nekoliko uzastopnih mjerenja ili iz dana u dan trebalo bi potaknuti sumnju na interferenciju krioglobulina.

Neposredno nakon proglašenja epidemije bolesti COVID-19 i Klinička bolnica Dubrava proglašena je Primarnim respiracijsko-intenzivističkim centrom (PRIC KBD). U vrlo kratkom roku bolnica se organizacijski prilagodila skrbi za pacijente oboljele od COVID-19, a sukladno tome Klinički zavod za laboratorijsku dijagnostiku prilagodio se analizi uzoraka oboljelih od COVID-19 sa svim predanalitičkim, analitičkim i poslijeanalitičkim procesima. Predanalitička faza laboratorijskog testiranja uključuje sve postupke prije analize uzorka te je zbog oskudnih podataka o zaraznosti uzoraka krvi, mokraće i ostalih tjelesnih tekućina bolesnika s COVID-19, prvi korak u reorganizaciji rada laboratorija bio upravo reguliran i definiran pristup u dekontaminaciji epruveta i spremnika bez utjecaja na stabilnost određivanih analita. Pritom je potrebno obratiti pozornost na vrlo učestale hiperkoagulabilne uzorke koji se analiziraju tek nakon opetovanog centrifugiranja. Povećane mjere opreza i zaštite provode se u likvorskoj dijagnostici, pogotovo pri brojenju stanica, zbog očekivano veće virulentnosti likvora. U analitičkom dijelu laboratorijskog procesa važno je osigurati kontinuiranu izradu svih analiza unatoč kompleksnosti izrade i izrazito promijenjenim laboratorijskim parametrima izvan granice linearnosti metode. Velik broj parametara se mijenja iznad uobičajeno definiranih kritičnih vrijednosti koje zahtijevaju neodgodivo obavještavanje kliničara za pravovremeno medicinsko zbrinjavanje bolesnika. Upravo su zato redefinirane kritične vrijednosti uz fokus samo na iznimno patološke ili promijenjene nalaze. Najčešće izvještavani rezultati jesu izrazito promijenjene vrijednosti acidobazičnog statusa i visoke koncentracije upalnih biljega poput C- reaktivnog proteina, interleukina 6, prokalcitonina, ali i feritina, koji odražavaju prekomjernu aktivaciju imunosnog sustava i citokinsku oluju. Upala i infekcija aktiviraju endotel, što rezultira aktivacijom trombocita i leukocita kao i promijenjenim antikoagulantnim i fibrinolitičkim mehanizmima koji se na laboratorijskim nalazima odražavaju kao povišene vrijednosti D-dimera, skraćeno vrijeme aktiviranoga parcijalnog tromboplastinskog vremena (APTV) i povišene vrijednosti aktivnosti protrombinskog vremena (PV). Period pandemije nametnuo je i uvođenje novih specijalističkih i visokodiferentnih pretraga neophodnih za kompletiranje laboratorijskog praćenja analiza COVID-19 poput SARS-CoV-2 PCR, SARS-CoV-2 IgG, interleukin 6, ali su uvedene i analize poput IgE, MPL (ekson10), M MYD88 L265P i Bence Jones protein imunofiksacijskom metodom, koje unaprjeđuju laboratorijsku dijagnostiku ostalih, vrlo prisutnih i ozbiljnih kliničkih dijagnoza.

BACKGROUND-AIM COVID-19 is an infectious disease caused by the virus called Severe acute respiratory syndrome Coronavirus-2 (SARS-CoV-2) that affects mainly the respiratory system, with mild symptoms in most cases, but that can also lead to critical conditions that require hospitalization. There is growing scientific evidence about the long- term effects of COVID-19 in a significant number of patients after the hospital stay. In our hospital, University Hospital Dubrava, these patients are monitored within a newly established Post-COVID Unit, and often referred to further respiratory rehabilitation regarding their decreased pulmonary function, among other complications. In this study we wanted to evaluate whether CYFRA 21-1 can be of prognostic importance in evaluation of lung injures in post-COVID patients. METHODS In our retrospective study we evaluated the levels of a well-known tumor biomarker Cytokeratin 19 fragment (CYFRA 21-1) in the serum of patients presented in the Post-COVID Unit a month or more after the end of hospitalization. For the quantitative determination of CYFRA 21-1 the Abbott’s ARCHITECT CYFRA 21-1 chemiluminescent microparticle immunoassay (CMIA) was used on the Architect i1000SR analyzer. Cytokeratin 19 is a part of the cytokeratin polypeptides family, support proteins that form the cytoskeleton of epithelial cells. CYFRA 21-1 is its serum-soluble fragment that can be detected in body fluids. It is quite frequently found in pulmonary tissue, in particular when a malignant lung tumor occurs. RESULTS CYFRA 21-1 levels were measured in 124 samples, of which 29 were above the decision limit used in our laboratory of 2.08 ng/mL, so we analyzed the available data for these 29 patients in more detail. There were 12 females and 17 males, from 82 to 49 years old, of which 7 with mild and 21 with severe symptoms and on oxygen therapy during hospitalization and unknown for one patient. None of them were on mechanical ventilation. In 26 patients a bilateral pneumonia was established during their hospital stay, and one patient was in home isolation. According to Thomas’s Clinical Laboratory Diagnostics, the decision level for patients with pulmonary disease is 3.3 ng/mL, and 12 of our patients were above this value, 10 of which with no previous history of pulmonary disease. CONCLUSION Having in mind that tumor markers are not used for diagnostic purposes, it is very unlikely that these patients are developing a lung carcinoma. However, this elevated CYFRA 21-1 levels may indicate a lung injury and therefore a greater need for future monitoring of their respiratory function.

BACKGROUND-AIM: Monocyte Distribution Width (MDW) is a novel hematology parameter, which represents a measure of change inthe monocyte size distribution. Morphological changes of monocytes in sepsis show greater heterogeneity of themonocyte population and thus a higher MDW value. MDW provides significant added value along with currently usedsepsis markers (WBC, PCT, CRP, IL-6) for early sepsis screening. The clinical MDW cut-off of 20.0 was establishedthrough a blinded prospective multi-center pilot study using K2-EDTA venous whole blood samples. According to thisstudy, MDW values >20.0 should raise suspicion that sepsis is present or will develop in patients within twelve hoursof the Emergency Department (ED) presentation. The type of EDTA salt may affect the accuracy of cell counting andsizing. The aim of our study was to evaluate the impact of Beckton-Dickinson (BD) blood collection tubes containingK2-EDTA and K3-EDTA as anticoagulants on MDW values. METHODS: Blood samples from 158 apparently healthy individuals undergoing annual health checkups were collected in K2-EDTA and simultaneously in K3-EDTA test tubes. All samples were analyzed for CBC with differential within 30 minafter collection on a Beckman Coulter Unicel DxH 900 hematology analyzer. Data were tested for normality usingthe Kolmogorov–Smirnov test. The significance of differences between samples was assessed by Wilcoxon's pairedtest. P < 0.05 was considered statistically significant. Data are presented as medians and interquartile ranges. Bland-Altman plotting was performed to assess the comparability of results. Statistical analysis was performed using MedCalc 14.8.1.0 statistical software. RESULTS: The study included 158 participants, age 30 (18–58), 91 (57, 6%) males. For K2-EDTA tubes median MDW value was 16, 25(15, 38–17, 44) and for K3-EDTA tubes 17, 11 (16, 18–18, 42). A statistically significant difference was found comparingMDW values in K2- EDTA and K3-EDTA (P

Introduction:Faecal calprotectin is a non- invasive biomarker of the inflammatory bowel disease diagnostics. The aim of this study was to verify the method (immunoturbidimetry) for calprotectin determination in a stool extract on the Olympus AU2700Plus automatic analyzer (Beckman Coulter Inc., Tokyo, Japan). Materials and methods: Verification was performed according to CSLI EP 15-A2 protocol. Commercial control samples at low (L1) and high (L2) levels (fCAL turbo, Bühlmann Laboratories AG, Switzerland) were used. The precision (CVp) and trueness (BIAS) were interpreted according to manufacturer’s criteria (20%). 33 examiner’s samples were used to compare immunoturbidimetry with ELISA method (fCAL ELISA, Bühlmann Laboratories AG, Schönenbuch, Switzerland) on the BEP 2000 Advance analyzer (Siemens Healtcare, Marburg, Germany). The results were analyzed by Passing-Bablok regression and Bland-Altman plot. Linearity was tested by serial dilutions (100%, 75%, 50%, 25%, 0%)in duplicate. Results:The obtained CVs were 6.3% (L1 ; target value 73.5 μg/g), 2.0% (L2 ; target value 245.5 μg/g) with mean CV of 4.2%. Trueness values were - 4.1% (L1), - 1.9% (L2) with mean BIAS of 3.0%. Passing-Bablok analysis yields the regression line Y = - 8.3083 + 0.9888X (95% CIfor the y-intercept - 26.6455 to - 1.4489 and for slope 0.9213 to 1.0523). Bland-Altman plot showed the average BIAS of 14.7%, which meets manufacturer’s criteria. The linearity test is within allowed deviation for each of the expected values in the dilution series. Conclusion:Determination of fecal calprotectin by the immunoturbidimetric method on the Olympus AU2700Plus analyzer meets the criteria for precision and trueness. The results of the comparison between the two methods indicate a constant error with no clinical significance, since immunochemical and immunological methods cannot be compared. Finally, the immunoturbidimetry is considered as acceptable method for routine application. The simple performance of the proposed method allows the clinician to obtain real-time information without testing delay caused by laboratory rationalization.

AIM: Sepsis is still the main cause of death in surgical intensive care unit (ICU) with continuously increasing incidence and mortality rate. Therefore, both early diagnosis and prognosis of sepsis are of great importance. In addition to procalcitonin (PCT), C-reactive protein (CRP), leukocytes an lactate, novel studies propose lipopolysaccharide-binding protein (LBP) as a sensitive marker for bacterial infection and possibly useful follow-up parameter of sepsis. The aim of this report is to investigate the additional clinical value of LBP to PCT, CRP, leukocytes and lactate in identification and outcome prediction of a patient with severe sepsis. PATIENT AND METHODS: A 74-year old male patient was admitted to ICU after major open-heart surgery. Seventeen days after surgery, severe sepsis with Pseudomonas aeruginosa infection was diagnosed. In the 10 following days before patients’ death concentrations of PCT (CLIA, Siemens Advia Centaur XP), CRP (immunoturbidimetry, Beckman Coulter AU680), leukocytes (optical count, Siemens Advia 2120i), lactate (photometry, Beckman Coulter AU680) and LBP (CLIA, Siemens Advia Centaur XP) were measured. The results were compared using the Reference Change Value (RCV), Z = 1.96. Obtained RCVs for PCT, CRP, leukocytes, lactate and LBP were 49%, 138%, 32%, 76% and 42%, respectively. These values were considered clinically significant. Sepsis-related Organ Failure Assesment (SOFA) score was calculated on the day of diagnosis. RESULTS: Measured values of all analytes on the first day of diagnosis were: CRP (138.8 mg/L), PCT (14.7 ng/mL), leukocytes (4.6x109/L), lactate (2.32 mmol/L) and LBP (45.9 μg/mL). Calculated SOFA score was 17 with estimation of mortality higher than 90%. The highest levels of CRP (298.6 mg/L), lactate (17.9 mmol/L) and leukocytes (16.6x109/L) were measured on the 3rd day showing clinically significant difference for leukocytes and lactate compared to the 1st day. CRP showed no significant difference within all 10 days. The highest values of PCT (32.85 ng/mL and 26.77 ng/mL) were on 2nd and 3rd day showing significant change and an expected continuous decrease in the following days, whereas LBP did not change significantly. CONCLUSION: Values for PCT, CRP and lactate remained within their patophysiological dynamics as was expected due to the patients’ condition and performed therapy. However, as leukocytes increased after the 5th day and LBP did not significantly decrease in all 10 days, these two parameters could be considered as predictors of fatal outcome in this case report. Also, as an increase of LBP was not observed, it is to presume that LBP starts to increase before PCT and CRP, and can help in identifying patients that are likely to develope severe sepsis days before other mentioned parameters. These results are in agreement with several studies but further research in this field is needed.

Introduction: The aim of this study is to present a case of the correction of nucleated red blood cell (NRBC)and white blood cell (WBC) count in a 32-year-old woman of South Asian origin with Hemoglobin E–beta thalassemia, followed in University Hospital Dubrava. Case report: The patient was followed for 16 months during regular monthly examinations in hematology clinic where her blood was collected in K3-EDTA coated tubes. Samples were first analyzed by Advia 2120i(Advia), followed by WBC count by flow cytometry(FCM ; CD45-ECD) and NRBC count/500WBCs in peripheral blood (PB) smear. Results obtained were divided into two groups: 1) Advia NRBC count Advia linearity range (112-295%) and 2) Advia NRBC count close to/beyond linearity range (≥295%). Within the group we compared WBC count between FCM and Advia auto-corrected (FCM/Advia) and between FCM and WBC count obtained by mathematical correction of uncorrected WBC count from Advia with NRBC count from PB smear (FCM/calculated). Data is expressed as medians and ranges. Differences between groups were compared using non-parametric Mann-Whitney test (P< 0.05 was considered statistically significant). Results: In group 1 WBC’s median (ranges): FCM 11(6, 1-15, 9), Advia 12, 2 (9, 3-16, 5), calculated 10, 5 (9, 2-16, 8), there was no statistically significant difference (FCM/Advia P=0, 310 ; FCM/calculated P=0, 402). In group 2 WBC’s median (ranges): FCM 10, 7 (6, 6-15, 6), Advia 14, 6 (11, 5-19, 9), calculated 11, 2 (8, 9-14, 6) there was no statistically significant difference for FCM/calculated (P=0, 805), but with difference being statistically significant for FCM/Advia (P=0, 026). Conclusion: Advia has algoritams which give reliable number of NRBCs in linear measurement range with an automatic correction of WBC and differential count, but NRBC count is not reliable above linearity range. In this case it is necessary to confirm NRBC count in PB smear. If NRBC count from Advia is above linearity range, it is necessary to report NRBC and WBC differential counts from PB smear and according to NRBC count from PB smear correct the uncorrected WBC count from Advia.