Analytical evaluation of the novel Mindray high sensitivity cardiac troponin I immunoassay on CL-1200i.

OBJECTIVES: The current study was designed to evaluate the analytical performance of the new Mindray highly sensitive cardiac troponin I (hs-cTnI) chemiluminescent immunoassay on Mindray CL-1200i, as a thorough validation of novel hs-cTnI methods is required before introduction into clinical practice. METHODS: The evaluation of the analytical performance of this hs-cTnI immunoassay encompassed the calculation of the limit of blank (LOB), limit of detection (LOD), functional sensitivity, imprecision, linearity, 99th percentile upper reference limit (URL) and concordance with another previously validated hs-cTnI chemiluminescent immunoassay. RESULTS: The LOB and LOD were 0.32 and 0.35 ng/L, whilst the functional sensitivity (expressed as cTnI value with <10 % imprecision), was 0.35 ng/L. The linearity was excellent throughout a wide range of clinically measurable values (r=1.00 between 0.8 and 9,726.9 ng/mL). The intra-assay, inter-assay and total imprecision were 1.1-1.3 %, 5.5-8.1 % and 5.6-8.2 %, respectively. The 99th percentile URL calculated using residual plasma from 246 ostensibly healthy blood donors was 9.2 ng/L (4.3 ng/L in women vs. 12.3 ng/L in men). The Spearman's correlation between Mindray hs-cTnI and Access hs-TnI was 0.97, with mean bias of 7.2 % (95 % CI, 2.6-11.9 %). CONCLUSIONS: Although we failed to confirm the very optimistic analytical characteristics previously reported for this method, our evaluation of the novel Mindray hs-cTnI immunoassay on CL-1200i demonstrated that the overall performance is comparable to that of other commercially available hs-cTnI techniques, making it a viable alternative to other methods.

Clinical assessment of SNIBE Maglumi SARS-CoV-2 antigen fully-automated chemiluminescent immunoassay.

OBJECTIVES: Given that SARS-CoV-2 antigen tests will represent a pillar for supporting or surrogating molecular testing in the endemic period, we report here the clinical performance of the new SNIBE Maglumi SARS-CoV-2 antigen fully-automated chemiluminescent immunoassay (MAG-CLIA SARS-CoV-2 Ag). METHODS: The study population consisted of 181 subjects (mean age 61 ± 21 years; 92 females) undergoing coronavirus disease 2019 (COVID-19) testing at the local diagnostic facility, from December 2022 to February 2023. Routine diagnostic practice involved the collection of a double nostril nasopharyngeal swab, analyzed in duplicate with SARS-CoV-2 antigen (MAG-CLIA SARS-CoV-2 Ag) and molecular (Altona Diagnostics RealStar SARS-CoV-2 RT-PCR Kit) tests. RESULTS: A significant Spearman's correlation was found between MAG-CLIA SARS-CoV-2 Ag and mean Ct values of SARS-CoV-2 E and S genes (r=-0.95; p<0.001). In all nasopharyngeal samples, the area under the curve (AUC) of MAG-CLIA SARS-CoV-2 Ag was 0.86 (95% CI, 0.81-0.90), with 0.71 sensitivity and 1.00 specificity at 7 ng/L cut-off, increasing to 0.98 (95% CI, 0.96-1.00) AUC and 0.96 sensitivity (with 0.97 specificity) in high viral load samples. When SARS-CoV-2 N protein concentration was replaced with raw instrumental readings (i.e., relative light units [RLU]), the AUC in all samples increased to 0.94. A RLU value of 945 was associated with 88.4% accuracy, 0.85 sensitivity, 0.95 specificity, 0.77 negative predictive value (NPV) and 0.97 positive predictive value (PPV), respectively. CONCLUSIONS: We found satisfactory analytical performance of MAG-CLIA SARS-CoV-2 Ag, which could be used as surrogate of molecular testing for identifying high viral load samples. Broadening the reportable range of values may generate even better performance.

Positivization time of a COVID-19 rapid antigen self-test predicts SARS-CoV-2 viral load: a proof of concept.

OBJECTIVES: This proof of concept study was aimed to validate the hypothesis that the time of positivization of SARS-CoV-2 self-performed rapid diagnostic tests (RDTs) may reflect the actual viral load in the specimen. METHODS: A SARS-CoV-2 positive sample with high viral load was diluted and concomitantly assayed with molecular assay (Xpert Xpress SARS-CoV-2) and RDT (COVID-VIRO ALL IN RDT). The (mean cycle threshold; Ct) values and RDT positivization times of these dilutions were plotted and interpolated by calculating the best fit. The parameters of this equation were then used for converting the positivization times into RDT-estimated SARS-CoV-2 Ct values in routine patient samples. RESULTS: The best fit between measured and RDT-estimated Ct values could be achieved with a 2-degree polynomial curve. The RDT-estimated Ct values exhibited high correlation (r=0.996) and excellent Deming fit (y=1.01 × x - 0.18) with measured Ct values. In 30 consecutive patients with positive RDT test, the correlation between RDT positivization time and measured Ct value was r=0.522 (p=0.003). The correlation of RDT-estimated and measured Ct values slightly improved to 0.577 (Deming fit: y=0.44 × x + 11.08), displaying a negligible bias (1.0; 95% CI, -0.2 to 2.2; p=0.105). Concordance of RDT-estimated and measured Ct values at the <20 cut-off was 80%, with 0.84 sensitivity and 0.73 specificity. CONCLUSIONS: This proof of concept study demonstrates the potential feasibility of using RDTs for garnering information on viral load in patients with acute SARS-CoV-2 infection.

Assessment of humoral and cellular immunity after bivalent BNT162b2 vaccination and potential association with reactogenicity.

OBJECTIVES: This study investigated the feasibility and clinical value of using a novel, automated and high-throughput SARS-CoV-2 Interferon Gamma Release Assay (IGRA), combined with total anti-SARS-CoV-2 antibodies assessment, for evaluating the immune response after bivalent BNT162b2 vaccination. METHODS: A cohort of healthcare workers, who already underwent primary vaccination and boosting with monovalent BNT162b2 vaccine, received a booster dose of the new BNT162b2 bivalent formulation. Blood samples were taken immediately before vaccination (T0) and 1 month afterwards (T1). Humoral and cellular immunity were assayed with Roche Elecsys Anti-SARS-CoV-2 and Roche Elecsys IGRA SARS-CoV-2, respectively. RESULTS: The study population consisted of 51 subjects (median age: 43 years; 51% females). Total anti-SARS-CoV-2 antibodies and IGRA SARS-CoV-2 values increased at T1 from 9,050 to 25,000 BAU/mL (p<0.001), and from 0.44 to 0.78 IU/mL (p=0.385), accounting for median increase of 2.0 and 1.6 folds, respectively. Increased T1 values of total anti-SARS-CoV-2 antibodies and IGRA SARS-CoV-2 were recorded in 100% and 68.6% subjects, respectively. In those with baseline values below the median, post-vaccine levels displayed larger increases of 3.3 and 5.1 folds for anti-SARS-CoV-2 total antibodies and IGRA SARS-CoV-2, respectively. The variation of total anti-SARS-CoV-2 antibodies was inversely associated with their T0 values (r=-0.97; p<0.001), whilst that of IGRA SARS-CoV-2 was inversely associated with its T0 value (r=-0.58; p<0.001). No other signifcant associations were found with demographical or clinical variables, including side effects. CONCLUSIONS: The bivalent BNT162b2 vaccine booster enhances humoral and cellular immunity against SARS-CoV-2, especially in recipients with lower baseline biological protection.

Effect of BNT162b2 booster dose on anti-SARS-CoV-2 spike trimeric IgG antibodies in seronegative individuals.

OBJECTIVES: We provide here an updated analysis of an ongoing serosurveillance study, presenting data on the effect of a third dose of Pfizer/BioNTech BNT162b2 vaccine on serum anti-SARS-CoV-2 IgG antibodies. METHODS: We tested baseline SARS-CoV-2 seronegative healthcare workers undergoing primary vaccination with the mRNA-based COVID-19 Comirnaty vaccine, followed by administration of homologous vaccine booster (third dose). Venous blood was collected before either dose of primary vaccination, at 1, 3 and 6 months afterwards, as well as before and 1 month after receiving the vaccine booster. The serum concentration of anti-SARS-CoV-2 IgG was assayed with DiaSorin Trimeric spike IgG immunoassay. RESULTS: The final study population included 53 SARS-CoV-2 seronegative healthcare workers (median age 46 years; 60% females). A first peak of anti-SARS-CoV-2 spike trimeric IgG values was reached 1 month after completing primary vaccination, after which the levels gradually declined until before receiving the vaccine booster. A second peak of anti-SARS-CoV-2 spike trimeric IgG concentration was observed 1 month after receiving the vaccine booster dose (8,700 kBAU/L), which was 39-fold higher than before receiving the vaccine booster (221 kBAU/L; p<0.001), but was also nearly threefold higher compared to values seen at the first peak (2,990 kBAU/L; p<0.001). The rate of subjects with protective anti-SARS-CoV-2 spike trimeric IgG values (i.e., >264 kBAU/L) increased from 47.2% to 100% after 1 month from vaccine booster. CONCLUSIONS: These results support current policies fostering COVID-19 vaccine boosters to reinforce humoral immunity against SARS-CoV-2.

Clinical performance of Fujirebio Lumipulse G SARSCoV- 2 Ag chemiluminescent immunoassay.

We evaluated the performance of Fujirebio Lumipulse G SARS-CoV-2 Ag chemiluminescent immunoassay. A nasopharyngeal swab was collected from 160 subjects and assayed simultaneously with Fujirebio Lumipulse G SARS-CoV-2 Ag and Altona Diagnostics RealStar SARS-CoV-2 RT-PCR assays. Using 0.60 pg/mL diagnostic threshold, Fujirebio Lumipulse G SARS-CoV-2 Ag displayed 0.88 area under the curve, 0.88 sensitivity and 0.75 specificity compared to molecular testing. The area under the curve increased to 1.00 after excluding samples with low viral load (i.e., cycle threshold values between 25-37). Thus, this chemiluminescent immunoassay could be used for rapid identification of many subjects with high nasopharyngeal SARS-CoV-2 viral load.

Comprehensive assessment of humoral response after Pfizer BNT162b2 mRNA Covid-19 vaccination: a three-case series.

OBJECTIVES: Since universal vaccination is a pillar against coronavirus disease 2019 (COVID-19), monitoring anti-SARS-CoV-2 neutralizing antibodies is essential for deciphering post-vaccination immune response. METHODS: Three healthcare workers received 30 µg BNT162b2 mRNA Covid-19 Pfizer Vaccine, followed by a second identical dose, 21 days afterwards. Venous blood was drawn at baseline and at serial intervals, up to 63 days afterwards, for assessing total immunoglobulins (Ig) anti-RBD (receptor binding domain), anti-S1/S2 and anti-RBD IgG, anti-RBD and anti-N/S1 IgM, and anti-S1 IgA. RESULTS: All subjects were SARS-CoV-2 seronegative at baseline. Total Ig anti-RBD, anti-S1/S2 and anti-RBD IgG levels increased between 91 and 368 folds until 21 days after the first vaccine dose, then reached a plateau. The levels raised further after the second dose (by ∼30-, ∼8- and ∼8-fold, respectively), peaking at day 35, but then slightly declining and stabilizing ∼50 days after the first vaccine dose. Anti-S1 IgA levels increased between 7 and 11 days after the first dose, slightly declined before the second dose, after which levels augmented by ∼24-fold from baseline. The anti-RBD and anti-N/S1 IgM kinetics were similar to that of anti-S1 IgA, though displaying substantially weaker increases and modest peaks, only 4- to 7-fold higher than baseline. Highly significant inter-correlation was noted between total Ig anti-RBD, anti-S1/S2 and anti-RBD IgG (all r=0.99), whilst other anti-SARS-CoV-2 antibodies displayed lower, though still significant, correlations. Serum spike protein concentration was undetectable at all-time points. CONCLUSIONS: BNT162b2 mRNA vaccination generates a robust humoral immune response, especially involving anti-SARS-Cov-2 IgG and IgA, magnified by the second vaccine dose.

Update on Patient Self-Testing with Portable and Wearable Devices: Advantages and Limitations.

Laboratory medicine has undergone a deep and multifaceted revolution in the course of human history, in both organizational and technical terms. Over the past century, there has been a growing recognition of the need to centralize numerous diagnostic activities, often similar or identical but located in different clinical departments, into a common environment (i.e., the medical laboratory service), followed by a progressive centralization of tests from smaller laboratories into larger diagnostic facilities. Nevertheless, the numerous technological advances that emerged at the beginning of the new millennium have helped to create a new testing culture characterized by a countervailing trend of decentralization of some tests closer to patients and caregivers. The forces that have driven this (centripetal) counter-revolution essentially include a few key concepts, namely "home testing", "portable or even wearable devices" and "remote patient monitoring". By their very nature, laboratory medical services and remote patient testing/monitoring are not contradictory, but may well coexist, with the choice of one or the other depending on the demographic and clinical characteristics of the patient, the type of analytical procedure and the logistics and local organization of the care system. Therefore, this article aims to provide a general overview of patient self-testing, with a particular focus on portable and wearable (including implantable) devices.

Vaccination time does not influence total anti-SARS-CoV-2 antibodies response.

We measured total anti-SARS-CoV-2 receptor-biding domain (RBD) antibodies in 249 healthcare workers (mean age: 44 ± 13 years; 151 women), who received the first dose of mRNA-based Comirnaty COVID-19 vaccine at different times of the day. Compared with the reference vaccination time point (i.e. <10:00h), vaccine injection at the following times of day elicited a comparable response (all p > 0.05). Under our experimental conditions, we can therefore exclude a possible influence of the timing of primary mRNA-based COVID-19 vaccination on the levels of total anti-SARS-CoV-2 antibodies.

Effect of syringe underfilling on the quality of venous blood gas analysis.

OBJECTIVES: There is limited information on the influence of collecting small amounts of blood on the quality of blood gas analysis. Therefore, the purpose of this study was to investigate the effects of different degrees of underfilling of syringes on test results of venous blood gas analysis. METHODS: Venous blood was collected by venipuncture from 19 healthcare workers in three 1.0 mL syringes for blood gas analysis, by manually aspirating different volumes of blood (i.e., 1.0, 0.5 and 0.25 mL). Routine blood gas analysis was then immediately performed with GEM Premier 5,000. The results of the two underfilled syringes were compared with those of the reference syringe filled with appropriate blood volume. RESULTS: The values of most assayed parameters did not differ significantly in the two underfilled syringes. Statistically significant variations were found for lactate, hematocrit and total hemoglobin, the values of which gradually increased as the fill volume diminished, as well as for sodium concentration, which decreased in both insufficiently filled blood gas syringes. The bias was clinically meaningful for lactate in syringe filled with 0.25 mL of blood, and for hematocrit, total hemoglobin and sodium in both syringes containing 0.5 and 0.25 mL of blood. CONCLUSIONS: Collection of smaller volumes of venous blood than the specified filling volume in blood gas syringes may have an effect on the quality of some test results, namely lactate, hematocrit, total hemoglobin and sodium. Specific indications must be given for standardizing the volume of blood to be collected within these syringes.

Effect of exogenous lipids contamination on blood gas analysis.

Objectives: The purpose of this study was to investigate the effects of contamination of venous blood with a lipid-containing solution on parameters measured by a modern blood gas analyzer. Methods: We collected venous blood from 17 healthcare workers (46 ± 11 years; 53 % women) into three blood gas syringes containing 0 , 5 and 10 % lipid-containing solution. Blood gas analysis was performed within 15 min from sample collection on GEM Premier 5000, while triglycerides and serum indices were assays on Roche COBAS C702. Results: Triglycerides concentration increased from 1.0 ± 0.3 mmol/L in the uncontaminated blood gas syringe, to 39.4 ± 7.8 and 65.3 ± 14.4 mmol/L (both p<0.001) in syringes with 5 and 10 % final lipid contamination. The lipemic and hemolysis indices increased accordingly. Statistically significant variation was noted for all analytes except hematocrit and COHb in the syringe with 5 % lipids, while only COHb did not vary in the syringe with 10 % lipids. Significant increases were observed from 5 % lipid contamination for pO2, SO2 and lactate, while the values of pH, pCO2, sodium, potassium, chloride, ionized calcium, glucose, hematocrit (10 % contamination), hemoglobin and MetHB decreased. All these changes except lactate and CoHb exceeded their relative performance specifications. Conclusions: Artifactual hyperlipidemia caused by contamination with exogenous lipids can have a clinically significant impact on blood gas analysis. Manufacturers of blood gas analyzers must be persuaded to develop new instruments equipped with serum indices.

Preanalytical Impact of Incomplete K2EDTA Blood Tube Filling in Molecular Biology Testing.

BACKGROUND AND AIMS: The aim of this study was to investigate the possible preanalytical effect of incomplete filling of blood tubes on molecular biology assays. MATERIALS AND METHODS: The study population consisted of 13 healthy volunteers from whom 11 mL of whole blood was collected and then distributed in different volumes (1.5, 3.0, and 6.0 mL, respectively) into three 6.0 mL spray-dried and evacuated K2EDTA blood tubes. Automated RNA extraction was performed using the Maxwell® CSC RNA Blood Kit. DNA was extracted with a MagCorePlusII, with concomitant measurement of glyceralde-hyde-3-phosphate dehydrogenase (GAPDH) gene expression. The nucleic acid concentration was calculated using the NanoDrop 1000 spectrophotometer, and purity was assessed using A260/280 and A260/230 absorbance ratios. RESULTS: The RNA concentration was higher in the tubes filled with 1.5 and 3.0 mL of blood than in the reference 6 mL filled tube. The RNA 260/280 and RNA 260/230 ratios did not differ significantly between the differently filled blood tubes. The DNA concentration remained constant in the differently filled tubes. Compared to the 6.0 mL reference filled tube, the 1.5 mL and 3.0 mL filled blood tubes displayed a lower DNA 260/280 nm ratio. The DNA 260/230 ratio did not differ significantly in any of the variously filled tubes. Compared to the 6.0 mL reference filled blood tube, the 1.5 mL and 3.0 mL filled blood tubes showed a significant increase in the GAPDHcycle threshold. CONCLUSIONS: Our results suggest that underfilling of K2EDTA blood tubes may be a modest but analytically significant source of bias in molecular biology testing.

Urine dipstick for screening plasma glucose and bilirubin in low resource settings: a proof-of-concept study.

Objectives: The purpose of this proof-of-concept study was to investigate whether a commercially available urine dipstick may provide potentially useful information for screening plasma glucose and bilirubin in human plasma samples. Methods: Glucose and bilirubin were assayed in 60 anonymized lithium-heparin residual plasma samples using the Roche COBAS 8000 or after pipetting 10 µL of plasma onto the pads of a commercial urine dipstick. Semiquantitative urine test results obtained with the dipstick were directly compared to paired test results obtained with COBAS. Results: Median plasma glucose values between COBAS and dipstick were slightly different (5.8 vs. 5.6 mmol/L; p=0.040), while no significant difference was found in bilirubin values between COBAS and dipstick (11.2 vs. 8.6 µmol/L; p=0.090). The Spearman's correlation between COBAS and dipstick was 0.83 (95% CI, 0.73-0.90; p<0.001) for plasma glucose and 0.78 (95% CI, 0.66-0.87; p<0.001) for plasma bilirubin, respectively. Cumulative agreement between COBAS and dipstick was high for both glucose (88%; kappa statistic statistics, 0.75; 95% CI, 0.58-0.92; p<0.001) and bilirubin (88%; kappa statistics, 0.76; 95% CI, 0.60-0.92; p<0.001). Conclusions: The results of this proof-of-concept study indicate that the commercial urine test strip used in our study provides acceptable performance for screening plasma glucose and bilirubin levels compared with reference laboratory assays.

Real-world assessment of the clinical performance of COVID-VIRO ALL IN rapid SARS-CoV-2 antigen test.

OBJECTIVES: Since the external validation of severe acute respiratory syndrome coronavirus 2 antigen rapid diagnostic tests (SARS-CoV-2 RDT-Ags) is a necessary requisite before they can be introduced into routine clinical practice, this study reports the results of a real-world assessment of the clinical performance of the new COVID-VIRO ALL IN device. METHODS: The study population consisted in 165 outpatients (median age: 43 years, range: 14-68 years; 66.1% females) who had paired nasal and nasopharyngeal samples collected upon hospital presentation. The samples were concomitantly tested with the AAZ-LMB COVID-VIRO ALL IN SARS-CoV-2 RDT-Ag and with Cepheid Xpert Xpress SARS-CoV-2 real-time reverse transcription polymerase chain reaction (RT-PCR). RESULTS: The number of subjects with positive RT-PCR results (i.e., mean Ct value <45) was 116 (70.3%), 109 (66.1%) and 86 (52.1%) with mean Ct values <37 and <30, respectively. In all RT-PCR positive samples, COVID-VIRO ALL IN displayed 78.8% agreement, 0.698 sensitivity, 1.000 specificity, 0.583 negative predictive value (NPV) and 1.000 positive predictive value (PPV) compared to RT-PCR. The median Ct value of samples testing positive with COVID-VIRO ALL IN was significantly lower than those testing negative (22.8 vs. 32.2; p<0.001). In samples with high viral load (i.e., Ct value <30), COVID-VIRO ALL IN displayed 92.1% agreement, 0.895 sensitivity, 0.949 specificity, 0.983 NPV and 0.951 PPV compared to RT-PCR. CONCLUSIONS: Although the diagnostic performance of COVID-VIRO ALL IN do not exactly match those of the manufacturer, its high NPV in high viral load samples would enable fast-track and rapid identification of highly contagious subjects.