Automated flow cytometry enables high performance point-of-care analysis of leukocyte phenotypes

Hoepelman

et al. 2020

JCM

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Nowadays, more trauma patients develop chronic critical illness (CCI), a state characterized by prolonged intensive care. Some of these CCI patients have disproportional difficulties to recover and suffer from recurrent infections, a syndrome described as the persistent inflammation, immunosuppression and catabolism syndrome (PICS). A total of 78 trauma patients with an ICU stay of ≥14 days (CCI patients) between 2007 and 2017 were retrospectively included. Within this group, PICS patients were identified through two ways: (1) their clinical course (≥3 infectious complications) and (2) by laboratory markers suggested in the literature (C-reactive protein (CRP) and lymphocytes), both in combination with evidence of increased catabolism. The incidence of PICS was 4.7 per 1000 multitrauma patients. The sensitivity and specificity of the laboratory markers was 44% and 73%, respectively. PICS patients had a longer hospital stay (median 83 vs. 40, p < 0.001) and required significantly more surgical interventions (median 13 vs. 3, p = 0.003) than other CCI patients. Thirteen PICS patients developed sepsis (72%) and 12 (67%) were readmitted at least once due to an infection. In conclusion, patients who develop PICS experience recurrent infectious complications that lead to prolonged hospitalization, many surgical procedures and frequent readmissions. Therefore, PICS forms a substantial burden on the patient and the hospital, despite its low incidence.

Section: Discussionmentioning

confidence: 99%

Persistent Inflammation, Immunosuppression and Catabolism Syndrome (PICS) after Polytrauma: A Rare Syndrome with Major Consequences

Hoepelman

et al. 2020

JCM

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“…All experiments were performed with whole blood using the fully automated AQUIOS CL® "Load & Go" flow cytometer (Beckman Coulter, Brea, CA, USA) at 30°C [32]. Firstly, the AQUIOS CL® automatically incubated whole blood with the double-labeled S. aureus bioparticles (end concentration of 10 × 10 6 /ml) and with antibodyfluorochrome conjugates for the neutrophil receptors CD16 (clone 3G8, PE labeled; Beckman Coulter) and CD62L (clone DREG56, ECD labeled; Beckman Coulter).…”

Section: Flow Cytometry Analysismentioning

confidence: 99%

“…Until now, however, flow analysis was too complex, operator-dependent, time-consuming, and laborious to be applicable in a clinical setting [28,29]. Recently, a flow cytometer became available that is able to prepare and analyze whole blood fast (< 25 min), highly reproducible, and fully automated [30][31][32]. Hence, we used such an approach to develop a new assay for neutrophil function (phagocytosis and phagosomal acidification) and neutrophil subsets that can be performed as a point-of-care test by any health care worker.…”

Section: Introductionmentioning

confidence: 99%

New automated analysis to monitor neutrophil function point-of-care in the intensive care unit after trauma

Fraiture

et al. 2020

ICMx

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Background: Patients often develop infectious complications after severe trauma. No biomarkers exist that enable early identification of patients who are at risk. Neutrophils are important immune cells that combat these infections by phagocytosis and killing of pathogens. Analysis of neutrophil function used to be laborious and was therefore not applicable in routine diagnostics. Hence, we developed a quick and pointof-care method to assess a critical part of neutrophil function, neutrophil phagosomal acidification. The aim of this study was to investigate whether this method was able to analyze neutrophil functionality in severely injured patients and whether a relation with the development of infectious complications was present. Results: Fifteen severely injured patients (median ISS of 33) were included, of whom 6 developed an infection between day 4 and day 9 after trauma. The injury severity score did not significantly differ between patients who developed an infection and patients who did not (p = 0.529). Patients who developed an infection showed increased acidification immediately after trauma (p = 0.006) and after 3 days (p = 0.026) and a decrease in the days thereafter to levels in the lower normal range. In contrast, patients who did not develop infectious complications showed high-normal acidification within the first days and increased tasset to identify patients at risk for infections after trauma and to monitor the inflammatory state of these trauma patients. Conclusion: Neutrophil function can be measured in the ICU setting by rapid point-of-care analysis of phagosomal acidification. This analysis differed between trauma patients who developed infectious complications and trauma patients who did not. Therefore, this assay might prove a valuable asset to identify patients at risk for infections after trauma and to monitor the inflammatory state of these trauma patients.

“…The AQUIOS CL R combines robotic automated sample preparation with analysis of cells using flow cytometry. 25 The AQUIOS CL The .lmd data files were exported from the AQUIOS CL R and imported into FlowJo R analysis software (Tree Star Inc., Ashland, OH).…”

Section: Flow Cytometry Analysismentioning

confidence: 99%

“…These limitations include a short time window-of-opportunity for analysis, because inflammatory cells are easily activated by ex vivo manipulation and flow cytometry requires a fully functional immunological laboratory with experienced laboratory personnel. 25,30 The technique of fully automated flow cytometry has become available that circumvents most of these limitations and is applicable in a field laboratory setting. 25,30 This now allows analysis of the immune system in athletes at the sports site.…”

mentioning

confidence: 99%

Analysis of human neutrophil phenotypes as biomarker to monitor exercise-induced immune changes

Journal of Leukocyte Biology

Bertinetto

et al. 2020

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The amplitude of the innate immune response reflects the degree of physiological stress imposed by exercise load. An optimal balance of exercise intensity and duration is essential for a balanced immune system and reduces the risk of dysfunction of the immune system. Therefore, it is hypothesized that neutrophils, as key players in the innate immune system, can be used as biomarker in detecting overtraining. The aim was to monitor the state of the innate immune system by phenotyping neutrophils during consecutive bouts of prolonged exercise. Study subjects were recruited from a cohort of walkers participating in a walking event on 3 consecutive days. Participants with immune deficiencies were excluded. Questionnaires to determine the physiological status of the participants were completed. Analysis of neutrophil receptor expression was done by a point-of-care fully automated flow cytometer. A total of 45 participants were recruited, of whom 39 participants were included for data analysis. Study participants had a median age of 64 (58-70) years. The absolute numbers CD16 dim /CD62L bright and CD16 bright /CD62L dim neutrophils were increased after the first 2 days of exercise followed by an adaptation/normalization after the third day. Participants with activated neutrophils (high CD11b expression) had an impaired physical feeling indicated by the participant on a lower visual analog scale compared to participants who did not have activated neutrophils (P = 0.017, P = 0.022). Consecutive days of prolonged exercise results in an initial systemic innate immune response, followed by normalization/adaptation. Increased neutrophil activation was associated with impaired physical feeling measured by a validated VAS score indicated by the participant. Fully automated point-of-care flow cytometry analysis of neutrophil phenotypes in a field laboratory might be a useful tool to monitor relevant differences in the systemic innate immune response in response to exercise. K E Y W O R D S innate immune system, neutrophil, prolonged walking, repetitive exercise 1 INTRODUCTION The immune system is very responsive to exercise, which leads to specific responses to acute and chronic exercise. 1-3 The amplitude of the Abbreviations: ANOVA, analysis of variance; BMI, body mass index; CD, cluster of differentiation; CR, complement receptor; DBP, diastolic blood pressure; HR, heart rate; IQR, interquartile range; MFI, median fluorescent intensity; SBP, systolic blood pressure; VAS, visual analog scale; WBC, white blood cell count. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.