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INTRODUCTION 3D-DIFF scattergram of the Mindray BC-6800 haematological analyser shows morphological abnormalities and lymphocyte cluster splitting related to the presence of reactive lymphocytes. This study aims to assess whether these cytographic changes are useful in detecting both activated and apoptotic lymphocytes, leading to an improvement in the laboratory diagnostic process of infectious mononucleosis. METHODS Two hundred three samples with modified shape and doubled lymphocyte cluster of DIFF scattergram (study group) were divided into two different subgroups: with and, respectively, without serological evidence of ongoing IM. Activated and apoptotic cells in peripheral blood were counted by light microscopy or gating in the instrumental dot plots. Values of apoptotic cells counted by microscopy were compared with those resulting from gating. RESULTS Samples with both shape change and doubled lymphocyte cluster had serological profiles according to the diagnosis of ongoing infectious mononucleosis. Blood smears review was positive for reactive lymphocytes in all 112 samples (100%). An underestimation of apoptotic cell count by light microscopy compared with the gating in the instrumental scatterplot was also observed (96 out of 112, 85.7%). CONCLUSION The additional lymphocyte cluster was significantly associated with activated and apoptotic lymphocytes in samples with serology suggesting ongoing infectious mononucleosis. Considering the significance of clue for infectious mononucleosis assigned to the apoptotic lymphocytes, a specific flag such as "apoptotic cells?" could be associate with the related cluster. Such a flag could be used for dedicated rules for smears review, thus increasing infectious mononucleosis detection in laboratories that do not usually practise instrumental cytograms observation.

SARS-CoV-2 virus is responsible for the current worldwide coronavirus disease 2019 (COVID-19) pandemic, infecting millions of people and causing hundreds of thousands of deaths. Understanding the antibody response to SARS-CoV-2 is crucial for the development of vaccines, therapeutics and public health interventions. However, lack of consistency in methods used to monitor antibody response to SARS-CoV-2 leaves some uncertainty in our fine understanding of the human antibody response mounted following SARS-CoV-2 infection. We developed a peptide-based enzyme-linked immunosorbent assay (ELISA) by selecting 7 synthetic peptides from the spike, membrane, and nucleocapsid protein sequences of SARS-CoV-2, which effectively detects the antibody response mounted by all COVID-19 convalescent tested. Strikingly, the assay shows a profound difference in antibody response among individual subjects, which may have a significant impact on disease severity. Together, our results define an efficient and specific serological assay to consistently measure the antibody response following SARS-CoV-2 infection, as well as help the design of vaccine and therapeuticals for prevention and treatment of COVID-19.

AimsThe presence of cold agglutinin in blood samples can cause a spontaneous agglutination of red blood cells (RBCs) when low temperature occurs. This phenomenon causes a spurious lowering of RBC count on the automated haematological analysers that are detected by incongruous values (≥370 g/L) of the mean cellular haemoglobi concentration (MCHC). A preheating at 37°C can remove the RBC agglutination generally resulting in a reliable count. It has been reported that the same result can be reached by using the optical reticulocyte (RET) channel of Sysmex analysers where the RBC count is not influenced by the presence of cold agglutinin. This study aims to evaluate these data in a larger population, with regard to environmental conditions on Sysmex analysers. We have also evaluated the influence of different thermal pretreatments on the RBC count.MethodsThis study was performed on 96 remnants of peripheral blood samples (48 with MCHC in normal range and 48 with MCHC>370 g/L) which have been analysed in different preanalytical conditions on the Sysmex analysers.ResultsA preheating of samples at 41°C for 1 min leads to a reversibility of the cold agglutination comparable to the one observed in the RET channel and yields better results compared with 37°C for 2 hours.ConclusionsNone of described procedures assure the complete cold agglutination reversibility in every case. Consequently, since the haematological analysers not yet provide reliable parameters to confirm the complete resolution of agglutination, further verification of RBC count accuracy needs to be performed.

Background The infectious mononucleosis (IM) includes clue elements, apoptotic and atypical lymphocytes. In IM, the evaluation of dot plot provided by Sysmex XN-1000 analyzer revealed a stretched lymphocytic cluster, white cell differential channel (WDF), on cytogram. Methods In this study, we analyzed 698 samples that include 39 IM, 76 chronic lymphoproliferative disorders, 25 nonclonal lymphocytosis, and 40 healthy donors. Five hundred eighteen samples with other diseases or interference were evaluated. The algorithm was validated on 40,000 files that were received from internal database of Sysmex-Dasit. Results The analysis of flow cytometry standard (FCS) files in WDF channel and presumed apoptotic lymphocytes counts on side scatter/forward scatter (SSC/FSC) and SSC/SFL (where SFL is side fluorescence) dot plot revealed excellent correlation among apoptotic cells on peripheral blood smear (R2 = 0.79 and 0.75). There was a variation of positional parameters in lymphocyte clusters WX, WY, and WZ. If WX-SSC > 500 and WY-SFL > 1,000 and WZ-FSC > 700, specificity equals to 99% and sensitivity equals to 100%. If nucleated red blood cell (NRBC)

Introduction Acute promyelocytic leukemia (APL) is a type of acute myeloid leukemia (AML) with a life-threatening coagulopathy. Once it is suspected, ATRA should be started. Appreciation of APL details is critical, but an experienced hematopathologist may not be available. We developed an algorithm, based on the parameters generated by automated blood cell counter ADVIA 2120i Siemens that can aid the diagnosis of APL. Methods All parameters in the algorithm were selected on the bases of the pathophysiology of the APL and the analyzer's technology. We used c1 cutoff: PLT 2; %saturated cells ≥ 1; % blasts > 4. Satisfying at least five of six cutoffs, we obtained an "APL criteria". It was tested on 247.209 raw-data from routine and emergency samples and on 124 raw-data from APL. We performed a multiparametric analyses and obtained definitive cutoffs (c2). It was validated on a new group of 51.002 raw-data from routine and emergency. Finally, it was tested on AML and ALL, MDS, MPN, oncologic samples, and hemolytic and megaloblastic anemia. Results The algorithm provided a high sensitivity (86.30%) and high specificity (99.83%) in APL with high or normal number of WBC and satisfactory results in APL with cytopenia (80%). The specificity was very high also in groups of hematological and nonhematological diseases. It can be used as an "APL criteria" to alert pathologists of the possible presence of APL. It together with instrumental and classical morphology may allow to reduce the time of diagnosis with further reduction in early death.

We evaluated an extended time in the microscopic review in samples in which the potential clinical information could be increased with respect to those that could be achieved with the usual laboratory methodologies. We used samples containing nucleated red blood cells in a small amount and cytopenic samples. For these purposes for each peripheral blood smear, the timing of eye-count differential was increased up to 20 min, regardless of the final number of cells which could be counted. In addition, an automated system for digital analysis of peripheral blood smears was employed and the number of cells counted was brought up to 1000 leukocytes. In both manual and automatic light microscopy extended observation, we obtained more diagnostic information in respect to those with routine or standard methods. Both automated and manual increase systems of the timing for microscopic review are useful tools to find diagnostic information that otherwise would be lost using normal and standard procedures. So, these methods should be used especially when there is a higher pre-test probability for discovery of pathological cells.

Summary Introduction B-chronic lymphocytic leukemia CLL, a neoplastic clonal disorder with monomorphous small B lymphocytes with scanty cytoplasm and clumped chromatin, can be morphologically differentiated in typical and atypical forms with different prognosis: Smudge cells (Gumprecht's shadows) are one of the well-known features of the typical CLL and are much less inconsistent in other different types CLPD. Abbott Cell-Dyn Sapphire uses the fluorescence after staining with the DNA fluorochrome propidium iodide for the measurement of nucleated red blood cells (NRBCs) and nonviable cells (FL3+ cell fraction): We have studied the possible correlation between presence and number of morphologically identifiable smudge cells on smears and the percentage of nonviable cells produced by Cell-Dyn Sapphire. Methods 305 blood samples from 224 patients with B-cell lymphoproliferative disorders and 40 healthy blood donors were analyzed by CBC performed by Cell-Dyn Sapphire, peripheral blood smear, and immunophenotype characterization. Result FL3+ fraction in CLPD directly correlated with the percentage of smudge cells and is significantly increased in patients with typical B-CLL. This phenomenon is much less evident in patients with atypical/mixed B-CLL and B-NHL. Conclusion In small laboratories without FCM and cytogenetic, smudge cells%, can be utilized as a preliminary diagnostic and prognostic tool in differential diagnosis of CLPD.

Premesse. I pitfalls sono trappole in grado di alterare i dati numerici citometrici e che bisogna conoscere bene al fine di evitarle. Il riconoscimento dei pitfalls e essenziale per il miglioramento dell’accuratezza e tutto il personale operante in Laboratorio che esegue e referta i test in urgenza deve essere in grado di riconoscerli. Il citofluorimetro Dasit “SYSMEX UF-1000i”, tramite il dimensionamento (FSC), la complessita cellulare (SSC) e la fluorescenza alta e bassa (FLH e FLL), e in grado di fornire citogrammi e distinguere le varie popolazioni cellulari urinarie, con notevole vantaggio nell’individuazione delle micro-ematurie.

Nel corso di uno screening preoperatorio viene individuato un paziente con notevole allungamento del tempo di tromboplastina parziale attivato (aPTT), normale tempo di protrombina (PT) e anamnesi negativa per precedenti episodi di sanguinamento. Vengono eseguiti l’aPTT sulla miscela 1:1 (plasma paziente:pool) e successivamente il dosaggio dei fattori di pertinenza della via intrinseca. La normalizzazione dell’aPTT nella miscela 1:1 (plasma paziente:pool) orienta per deficit di uno o piu fattori della “via intrinseca”, tuttavia la valutazione dei fattori XII, XI, IX e VIII non evidenzia carenze. Viene pertanto ipotizzato un deficit di uno degli altri fattori della fase di contatto (pre-callicreina, chininogeno ad alto peso molecolare). La mancata normalizzazione dell’aPTT nella miscela 1:100 (pool:plasma paziente) consente di escludere la carenza di pre-callicreina (PK), rendendo piu consistente l’ipotesi di un deficit di chininogeno ad alto peso molecolare (HMWK). In assenza del saggio immunologico, viene eseguito l’aPTT sulla miscela costituita dal plasma del paziente e dal 12,5% di plasma normale. La correzione dell’aPTT consente di confermare il sospetto di deficit di HMWK. Il paziente viene pertanto regolarmente sottoposto a intervento chirurgico senza complicanze emorragiche. Anche in assenza di test specialistici e costosi e possibile giungere a diagnosi di patologie piuttosto rare, utilizzando metodologie diagnostiche economiche e disponibili in tutti i Laboratori, quali l’attenta analisi dell’anamnesi in combinazione con la valutazione dei risultati dell’aPTT e del test di miscela.

Early coagulation support is essential in massively bleeding patients. A Coagulation Support Algorithm (CSA), integrating rapid TEG (r-TEG) and functional fibrinogen TEG (ff-TEG) could shorten the time to a tailored treatment (Figure ​(Figure11). Figure 1 Coagulation Support Algorithm. Methods A retrospective comparison of the time to available TEG and Standard Coagulation Tests (SCT: INR, aPTTr, fibrinogen level) results in two groups of bleeding and coagulopathic patients using citrate kaolin-TEG (k-TEG) or the CSA protocol (r-TEG/ff-TEG). Statistical analysis was performed with Student's t test for unpaired samples.