Automated, Point-of-Care mobile flow cytometry: Bringing the laboratory to the sample.

Background: Innate effector cells are very responsive to infectious and inflammatory cues found in damaged and inflamed tissues. Their activation is a potential target to assess the state of the immune system. Unfortunately, these cells are very susceptible for ex-vivo activation, hampering accurate interpretation of flow cytometry data. Whether a brief window exists before ex-vivo activation starts to occur is currently unknown. Aims: 1) This study extensively investigated ex-vivo activation of innate effector cells over time. 2) We tested the feasibility of applying a mobile, automated, flow cytometry laboratory for out-of-hospital Point-of-Care analyses to minimize ex-vivo activation bias. Methods: 1) Ex-vivo neutrophil, eosinophil and monocyte activation in a blood collection tube over time and the reactivity to a formyl-peptide was investigated in a healthy cohort. 2) To facilitate fast, out-of-hospital analysis, application of the mobile flow cytometry was tested by placing an automated flow cytometer into a van. The stability of the setup was assessed by repetitively measuring laser alignment and fluorescence verification beads. Findings: 1) Immediately after venipuncture activation marker expression on neutrophils, eosinophils and monocyte subsets started to change in a time-dependent manner. 2) The mobile flow cytometry laboratory travelled over 3000 km, performing measurements at 19 locations with a median single-person-set-up time of 14 min. The laser alignment and fluorescence were stable during all experiments. Conclusions: Accurate flow data of innate immune cells are only obtained when ex-vivo activation is kept to minimum. The use of a mobile, fast, automated, flow cytometry laboratory for out-of-hospital Point-of-Care analyses provides new investigational and diagnostic possibilities outside major hospital flow cytometry laboratories.

Neutrophil and Eosinophil Responses Remain Abnormal for Several Months in Primary Care Patients With COVID-19 Disease.

Introduction: Neutrophil and eosinophil activation and its relation to disease severity has been understudied in primary care patients with COVID-19. In this study, we investigated whether the neutrophil and eosinophil compartment were affected in primary care patients with COVID-19. Methods: COVID-19 patients, aged ≥ 40 years with cardiovascular comorbidity presenting to the general practitioner with substantial symptoms, partaking in the COVIDSat@Home study between January and April 2021, were included. Blood was drawn during and 3 to 6 months after active COVID-19 disease and analyzed by automated flow cytometry, before and after stimulation with a formyl-peptide (fNLF). Mature neutrophil and eosinophil markers at both time points were compared to healthy controls. A questionnaire was conducted on disease symptoms during and 3 to 6 months after COVID-19 disease. Results: The blood of 18 COVID-19 patients and 34 healthy controls was analyzed. During active COVID-19 disease, neutrophils showed reduced CD10 (p = 0.0360), increased CD11b (p = 0.0002) and decreased CD62L expression (p < 0.0001) compared to healthy controls. During active COVID-19 disease, fNLF stimulated neutrophils showed decreased CD10 levels (p < 0.0001). Three to six months after COVID-19 disease, unstimulated neutrophils showed lowered CD62L expression (p = 0.0003) and stimulated neutrophils had decreased CD10 expression (p = 0.0483) compared to healthy controls. Both (un)stimulated CD10 levels increased 3 to 6 months after active disease (p = 0.0120 and p < 0.0001, respectively) compared to during active disease. Eosinophil blood counts were reduced during active COVID-19 disease and increased 3 to 6 months after infection (p < 0.0001). During active COVID-19, eosinophils showed increased unstimulated CD11b (p = 0.0139) and decreased (un)stimulated CD62L expression (p = 0.0036 and p = 0.0156, respectively) compared to healthy controls. Three to six months after COVID-19 disease, (un)stimulated eosinophil CD62L expression was decreased (p = 0.0148 and p = 0.0063, respectively) and the percentage of CD11bbright cells was increased (p = 0.0083 and p = 0.0307, respectively) compared to healthy controls. Conclusion: Automated flow cytometry analysis reveals specific mature neutrophil and eosinophil activation patterns in primary care patients with COVID-19 disease, during and 3 to 6 months after active disease. This suggests that the neutrophil and eosinophil compartment are long-term affected by COVID-19 in primary care patients. This indicates that these compartments may be involved in the pathogenesis of long COVID.

The relevance of geriatric assessment for older patients receiving palliative chemotherapy.

OBJECTIVES: No tools accurately discriminate between older patients who are fit and those who are frail to tolerate systemic palliative treatment. This study evaluates whether domains of geriatric assessment (GA) are associated with increased risk of chemotherapy intolerance in patients who were considered fit to start palliative chemotherapy after clinical evaluation by their treating clinician. MATERIALS AND METHODS: This prospective multicenter study included patients ≥70 years who started first line palliative systemic treatment. Before treatment initiation, patients completed GA including Activities of Daily Life (ADL), Instrumental Activities of Daily Life (IADL), Mini-Mental State Examination (MMSE), Mini Nutritional Assessment (MNA), Geriatric Depression Scale (GDS-15) and the Timed Up and Go Test (TUGT). Primary endpoint was treatment modification, defined as inability to complete the first three sessions of systemic treatment as planned. Secondary endpoint was treatment related toxicity ≥ grade 3 (CTCAE Version 4). The association between GA and endpoints were assessed using univariable and multivariable logistic regression analysis. RESULTS: Ninety-nine patients with median age of 77 (+/- 8) years underwent GA. 48% of the patients required treatment modification and grade 3 toxicity occurred in 53% of patients. One or more geriatric impairments were present in 71% of patients and 32% of patients were frail in two or more domains. Only TUGT was associated with treatment modifications (OR 2.9 [95% CI 1.3-6.5]) and grade 3 toxicities (OR 2.8 [95% CI 1.2-6.3]). CONCLUSION: Frailty was common in older patients who were considered fit to receive palliative chemotherapy. Treatment modification was necessary in half of the patients. Only TUGT was significantly associated with treatment modifications and grade 3 chemotherapy toxicities.