Particle-enhanced Turbidimetric Immunoassay Research Articles

#996 Validation of peritoneal NGAL as a biomarker for peritonitis: a comparison between laboratory-base method and rapid stick test

Abstract Background and Aims Neutrophil gelatinase-associated lipocalin (NGAL) is a protein involved in iron homeostasis, inflammation, and fibrosis. Initially discovered in human granulocytes, NGAL is found in various tissues. In renal contexts, NGAL's small size (23-25 kDa) and positive charge aid glomerular filtration and absorption in the proximal tubule. Notably, elevated NGAL levels were identified in peritoneal fluids during peritonitis in 2009, establishing it as a biomarker for early Peritoneal Dialysis (PD)-associated peritonitis diagnosis for values more than 100 μg/L. While our medical centre currently relies on the lab-based NGAL assay (NGALlab), advancements introduce the NGAL dipstick test (NGALds), offering a rapid and semi-quantitative colorimetric assessment. This study compares NGALds with NGALlab in clinical practice, assessing NGALds's potential for early peritonitis detection in PD patients' peritoneal fluid. Method This study involves a validation of diagnostic accuracy conducted through a retrospective analysis of parallel samples obtained from peritoneal effluents. Samples were collected from both healthy patients during routine analysis and from cases with suspected or confirmed peritonitis. Both NGALlab and NGALds analyses were conducted on each sample. The analysis aimed to evaluate the performance of the new NGALds by comparing it with the established NGALlab. Peritoneal NGALlab quantification employed the standard BioPorto test (BioPorto Diagnostics, Hellerup, Denmark), utilizing a particle-enhanced turbidimetric immunoassay. The novel NGALds is a rapid semi-quantitative assay that measures NGAL levels in biological fluids through colourimetric test strips, utilizing an antibody sandwich lateral flow dipstick test. NGAL values were categorized into colour-coded levels: 25 μg/L, 50 μg/L, 100 μg/L, 150 μg/L, 300 μg/L, and 600 μg/L. Statistical analyses were performed using SPSS Statistics 26.0 software. Spearman rank correlation coefficients (Rs) assessed categorical groups of NGAL values for peritoneal NGALds and NGALlab tests. A graphic comparison of these two tests was presented through a Bland-Altman plot Results Out of 301 peritoneal effluent samples, 156 were diagnosed with suspected peritonitis by ISPD Criteria. As Table 1 showed, NGALds showed a strong correlation with NGALlab categories (Rs = 0.876, P = 10−96), successfully diagnosing 96% (150 out of 156) peritonitis cases. Discrepancies between NGALlab and NGALds, visualized in the Bland-Altman plot (Fig. 1), were most notable when NGAL values exceeded the 600 μg/L threshold. However, this difference doesn't impact clinical significance, as high NGAL values remain diagnostically for peritonitis with both methods. Conclusion In conclusion, our study underscores a significant correlation between NGAL values measured by the laboratory method (NGALlab) and the NGAL dipstick test (NGALds). Importantly, NGALds exhibited a strong capability, correctly identifying 96% of peritonitis cases determined by NGALlab. Beyond statistical correlation, the NGAL dipstick emerges as a valid alternative for swift evaluation in patients presenting symptoms suggestive of peritonitis. Its ease of use, rapidity, and semi-quantitative colorimetric assessment make NGALds a patient-friendly tool. This user-friendly characteristic holds immense potential for streamlined clinical workflows, allowing for prompt and efficient peritonitis diagnosis.

#1436 Donor plasma cystatin C, but not creatinine, is associated with measured GFR at one year after kidney transplantation

Abstract Background and Aims Although widely used during deceased kidney donor selection, donor plasma creatinine has limited capacity to predict future recipient kidney graft function. Cystatin C is a muscle-mass independent kidney function marker that could serve as alternative for this purpose. However, the association between donor cystatin C and subsequent recipient graft function remains unclear. We aim to assess the association of pre-donation cystatin C concentrations with recipient iothalamate-measured GFR (mGFR) one year after kidney transplantation and compare it to the association of donor creatinine concentrations. Method All deceased kidney donors, within the affiliated donation region of the University Medical Center Groningen (UMCG), who donated at least one kidney to a kidney transplant recipient (KTR) affiliated to the same hospital during a period of 15 years (2004-2019) were included. We excluded all subjects with missing data on donor plasma cystatin C or recipient mGFR at 1 year. Donor plasma creatinine concentrations were measured using an enzymatic assay, while donor cystatin C concentrations were measured using a validated particle-enhanced turbidimetric immunoassay (Roche). Associations of donor plasma creatinine and cystatin C concentrations with recipients’ mGFR one year after transplant were assessed using linear regression. Results A total of 53 deceased kidney donors (55% male, age 47 ± 18 years, 47% donation after circulatory death) donated a kidney to 53 kidney transplant recipients (55% male, age 57 ± 12 years). Median pre-donation plasma creatinine and cystatin C concentrations were 64 [55 to 82] μmol/L and 0.64 [0.51 to 0.82] mg/L respectively. Mean recipient mGFR one year after transplantation was 52.7 ± 17.1 ml/min. In univariable linear regression, higher donor cystatin C was significantly associated with lower mGFR (β −0.42, 95% CI: −0.67 to −0.16, p = 0.002), while donor creatinine was not associated (β −0.03 95% CI: −0.32 to 0.25, p = 0.81). The association between donor cystatin C and recipients’ mGFR remained independent of adjustment for potential confounders, including donor sex and age, t (p = 0.008). Conclusion Higher donor cystatin C, but not creatinine, is associated with lower measured GFR one year after kidney transplant. Consequently, donor cystatin C could be valuable for inclusion in the pre-transplant evaluation of deceased kidney donors.

#758 NGAL dipstick for peritonitis: bridging the gap between rapid assessment and clinical efficiency in peritoneal dialysis

Abstract Background and Aims Neutrophil gelatinase-associated lipocalin (NGAL) is a versatile protein known for its multifunctional roles, extending beyond its conventional use as a biomarker. It actively participates in crucial biological processes such as inflammation, proliferation, and migration. This study seeks to evaluate NGALds, a recently developed dipstick test, in comparison to the established laboratory method (NGALlab) for the early detection of peritonitis related to peritoneal dialysis (PD). The focus of the investigation lies in analyzing peritoneal fluid to develop a rapid and effective diagnostic tool, aiming to enhance the management of peritonitis. Method A retrospective analysis was undertaken to validate diagnostic accuracy, utilizing parallel samples obtained from peritoneal effluents. Samples were obtained from both PD stable patients undergoing routine analysis and from PD patients with suspected or confirmed peritonitis. The assessment of NGALds involved a direct comparison with the established NGALlab. Peritoneal NGALlab quantification was conducted using the BioPorto test, employing a particle-enhanced turbidimetric immunoassay. The NGALds, a rapid semi-quantitative assay, utilized colorimetric test strips and an antibody sandwich lateral flow dipstick test. The greatest value of NGAL detected by dipstick is 600 μg/L. Statistical analyses, including Spearman rank correlation coefficients (Rs), were performed with SPSS Statistics 26.0 software. Additionally, ROC analysis was carried out using white cell count and percentage of Neutrophils in peritoneal fluids as the gold standard. Results Among the 301 peritoneal effluent samples analyzed, 156 were diagnosed with suspected peritonitis using NGALlab and ISPD criteria. Each sample underwent analyses with both NGALlab and NGALds. As depicted in Fig. 1, NGALds exhibited a robust correlation with NGALlab categories (Rs = 0.876, P < 0.001), accurately identifying 96% (150 out of 156) of peritonitis cases. Discrepancies between NGALlab and NGALds were most evident when NGAL values exceeded the 600 μg/L threshold. Nevertheless, these variations did not impact clinical significance, as both methods consistently diagnosed elevated NGAL values indicative of peritonitis. After ROC analysis, which yielded an AUC of 0.82 (p < 0.001) (Fig. 2), the Youden Index was employed to identify a threshold value of 100 μg/L for the diagnosis of suspected peritonitis. Importantly, the application of this threshold to NGALds demonstrated a positive predictive value of 0.64 and a negative predictive value of 0.87. Conclusion In conclusion, our study reinforces the significant correlation between NGAL values obtained through the laboratory standard (NGALlab) and the novel NGAL dipstick test (NGALds). Notably, NGALds exhibited high efficacy, accurately identifying 96% of peritonitis cases diagnosed by NGALlab. This robust performance positions the NGAL dipstick as a valuable and accessible alternative for rapid assessment in patients exhibiting symptoms indicative of peritonitis. The user-friendly attributes of NGALds not only make it a practical diagnostic tool but also a patient-centric one. Furthermore, the high negative predictive value provides confidence in considering NGALds as a frontline diagnostic tool for suspected peritonitis. The ease of use and prompt results open a promising avenue for improving clinical workflows, offering an opportunity for expedited and efficient peritonitis diagnosis. For instance, NGALds could find utility in centres where NGAL is unavailable in the laboratory or at the patient's home.

Serum Biomarker Analysis in Pediatric ADHD: Implications of Homocysteine, Vitamin B12, Vitamin D, Ferritin, and Iron Levels.

The current diagnosis of attention deficit hyperactivity disorder (ADHD) is based on history, clinical observation, and behavioral tests. There is a high demand to find biomarkers for the diagnosis of ADHD. The aim of this study is to analyze the serum profiles of several biomarkers, including homocysteine (Hcy), vitamin B12, vitamin D, ferritin, and iron, in a cohort of 133 male subjects (6.5-12.5 years), including 67 individuals with an ADHD diagnosis based on DSM-V criteria and 66 age-matched healthy boys (healthy controls, HC). Assessments for ADHD included the Iowa Conners' Teacher Rating Scale (CPRS) and the ADHDT test, as well as cognitive assessments using the Wechsler Intelligence Scale for Children-Revised (WISC-R) and the TROG-2 language comprehension test. Hcy and iron were quantified using spectrophotometry, while vitamin B12 and total 25-hydroxy vitamin D levels were determined using an electrochemiluminescence immunoassay (ECLIA) and ferritin was measured using a particle-enhanced immunoturbidimetric assay. The results showed significantly increased Hcy levels and decreased vitamin B12 levels in ADHD patients compared to HCs. Multiple logistic regression analysis indicated that Hcy is a potential prognostic indicator for ADHD. These results suggest that elevated homocysteine and decreased vitamin B12 may serve as markers for the diagnosis and prognosis of ADHD.

Establishing reference intervals for soluble urokinase plasminogen activator receptor in Northern European adults

ObjectivesSoluble urokinase plasminogen activator receptor (suPAR) may have untapped potential in clinical diagnostics. Previous studies determined reference intervals using an enzyme-linked immunoassay, but there is a need for reference intervals using a faster assay if the analysis is to be used in emergency medicine. The current study aims to determine reference intervals for suPAR using a fully automated particle-enhanced turbidimetric immunoassay (PETIA) according to the Clinical and Laboratory Standards Institute guideline A28-A3c. Design and methodsBlood samples were prospectively collected from Danish blood donors. Plasma suPAR was analyzed on the cobas 8000 module c502 in an open channel using a PETIA. Sex-partitioned reference intervals were determined using a parametric quantile approach. ResultsThe study included 241 participants—123 females and 118 males. The common reference interval for suPAR was 1.56–4.11 ng/mL (95% confidence intervals (CI) for the lower and upper limits were 1.56–1.63 and 3.81–4.47, respectively). The reference interval for females was 1.59–4.65 ng/mL (95% CIs 1.48–1.70 and 4.09–5.48, respectively) and for males, 1.56–3.59 ng/mL (95% CIs 1.47–1.65 and 3.31–3.93, respectively). ConclusionsOur results support using sex-partitioned reference intervals for suPAR and provide a basis for future studies using the PETIA method.

Three-minute nebulization of gentamicin in healthy dogs results in therapeutic concentrations in bronchoalveolar lavage fluid while remaining below the toxic range values in blood.

To determine and compare the concentration of gentamicin in the lower airways and serum of healthy spontaneously breathing dogs after nebulization with 5% undiluted gentamicin during 3 versus 10 minutes. 10 healthy experimental Beagles. This was a prospective crossover study. A standardized bronchoalveolar lavage (BAL) procedure was performed in each dog after 1 week of administration of each of 2 different gentamicin nebulization protocols separated by a 1-week washout period. The 2 protocols consisted of nebulization of 5% undiluted gentamicin (50 mg/mL) twice daily either during 10 minutes per session (± 95 mg; 10-minute protocol) or 3 minutes per session (± 28 mg; 3-minute protocol). BAL fluid (BALF) was obtained under general anesthesia using a bronchoscope within 15 minutes after administration of the last nebulization. Blood was collected within 5 minutes after BALF collection. BALF and serum gentamicin concentrations were determined by particle-enhanced turbidimetric inhibition immunoassay. Concentrations between protocols were compared using a paired t test. Both BALF and serum gentamicin concentrations were higher after the 10-minute protocol compared with the 3-minute protocol (mean ± SD: 2.41 ± 0.87 mg/L vs 1.25 ± 0.31 mg/L, P = .001; and 1.02 ± 0.59 mg/L vs 0.31 ± 0.24 mg/L, P < .0001 in BALF and serum, respectively), while the BALF-to-serum ratio did not differ between the protocols (3.75 [1.37 to 5.75] (median [IQR]) in the 3-minute protocol vs 2.48 [2.02 to 2.67] in the 10-minute protocol; P = .754). A 3-minute nebulization of gentamicin seems to achieve sufficient concentrations of gentamicin in the BALF to have good efficacy against aminoglycoside-sensitive bacteria while remaining below the toxic range values in blood.

Validation of rapid fecal calprotectin assay using particle enhanced turbidimetric immunoassay for inflammatory bowel disease.

Fecal calprotectin (FC) is a promising biomarker for diagnosis and treatment of inflammatory bowel disease, ulcerative colitis (UC), and Crohn's disease. An enzyme immunoassay (EIA) is widely used for FC detection, though the considerable lag time, up to several days, causes clinical management delay. This study was performed to examine the new rapid kit fCAL-turbo, which is based on a particle-enhanced turbidimetric immunoassay (15 min), by comparing FC values with other EIAs (EliA, PhiCal, Bühlmann) and endoscopic scores. Using 94 samples, fCAL-turbo showed strong significant positive correlations with the other kits (Spearman's r = 0.9178-0.9886). Of 74 UC patients, 69 underwent an endoscopy and fCAL-turbo reflected endoscopic activity with a moderate correlation with Mayo endoscopic subscore (MES) (r = 0.6945, others r = 0.6682-0.7013). Receiver operating characteristic analyses based on MES 0 versus 1-3 showed a similar efficacy as compared to the other kits (cut-off and area under the curve: 89.70 µg/g and 0.8592, respectively, others 62.35-138.4 µg/g and 0.8280-0.8611, respectively). Furthermore, multiple regression analysis confirmed that fCAL-turbo results significantly contributed to prediction of MES 0 with a higher t-value as compared to the other biomarkers. fCAL-turbo showed strong correlations with the other kits and also demonstrated excellent performance for predicting endoscopic remission of UC.

Significance of Inflammatory Markers; C-reactive Protein, Hyperuricemia, and Albuminuria in type 2 Diabetes Patients

Objective: To determine the significance of inflammatory markers, including C-reactive protein, hyperuricemia, and albuminuria, in individuals with type 2 diabetes.&#x0D; Methodology: This cross-sectional analytical study was done at Diabetic Clinic and Nephrology Department of Akbar Niazi Teaching Hospital, Islamabad between Jan 2023 and July 2023. Total 150 consecutive patients aged between 35 and 60 years with type 2 diabetes were enrolled in study. Participants were divided in two groups on the basis of C-reactive protein and uric acid levels. Group-A comprised individuals with type 2 diabetes and C-reactive protein levels, while Group-B included individuals with type 2 diabetes and uric acid levels. Every patient underwent necessary clinical and laboratory assessments. The measurement of C-reactive protein was conducted through a particle-enhanced immunoturbidimetric assay.&#x0D; Results: The study population demonstrates an average age of 49.41±7.66 years. In linear regression analysis, both C-reactive protein and uric acid emerged as independently predictor the albumin to creatinine ratio include (?=0.20, 95% CI 0.1-0.30, and p=0.001 for C-reactive protein, and ?=0.20, 95% CI 0.1-0.30, and p=0.001 for uric acid). In logistic regression, the albuminuria odd for C-reactive protein and uric acid groups was 4.05 (95% CI 2.60-9.15, p=0.001).&#x0D; Conclusion: In individuals with type 2 diabetes, there was an association observed between albuminuria and heightened levels of C-reactive protein and elevated uric acid levels. Additionally, there was an interactive effect on albuminuria observed between C-reactive protein and uric acid.&#x0D;

Macrolide-resistant Mycoplasma pneumoniae in an Iranian Pediatric Sample with Community-Acquired Pneumonia

Background: To the best of our knowledge, the prevalence of macrolide-resistant Mycoplasma pneumoniae (MRM) in Iranian children has not been investigated. Objectives: The present study aimed to evaluate the prevalence of MRM in Iranian children with community-acquired pneumonia (CAP). Methods: A total of 222 children with CAP, aged 3 - 15 years, who were hospitalized in 10 different children's hospitals, were enrolled in this study. Mycoplasmas were detected using the polymerase chain reaction (PCR) assay. The severity of CAP was evaluated according to the Infectious Diseases Society of America (IDSA) guidelines. The level of C-reactive protein (CRP) was also measured by the particle-enhanced turbidimetric immunoassay. Additionally, the chest X-rays of children with CAP were recorded and sent to a radiologist for further evaluation. Results: Twenty-one children (9.4%) diagnosed with CAP also had M. pneumoniae infection, 17 (77.27%) of whom were positive for A2063G transition and high-level macrolide resistance. The severity of CAP (P ≥ 0.99), CRP level (0.07), and chest X-ray changes (P = 0.08) were not significantly different between children with MRM pneumonia and those with macrolide-susceptible M. pneumoniae. Conclusions: The prevalence of high-level MRM pneumonia in children is high in Iran, similar to other Asian countries. However, this type of Mycoplasma infection was not associated with the severity of CAP and did not have significant effects on chest X-ray (CXR) changes or the CRP level in the patients.

A novel serum calprotectin (MRP8/14) particle-enhanced immuno-turbidimetric assay (sCAL turbo) helps to differentiate systemic juvenile idiopathic arthritis from other diseases in routine clinical laboratory settings

BackgroundDifferential diagnosis in children with signs of unprovoked inflammation can be challenging. In particular, differentiating systemic juvenile idiopathic arthritis (SJIA) from other diagnoses is difficult. We have recently validated the complex of myeloid-related proteins 8/14 (MRP8/14, also known as S100A8/A9 complex or serum calprotectin) as a helpful biomarker supporting the diagnosis of SJIA. The results were subsequently confirmed with a commercial ELISA. However, further optimization of the analytical technology is important to ensure its feasibility for large-scale use in routine laboratory settings.MethodsTo evaluate the accuracy in identifying children with SJIA, the performance of a particle-enhanced immuno-turbidimetric assay for serum calprotectin (sCAL turbo) on an automated laboratory instrument was analyzed. Samples from 615 children were available with the diagnoses SJIA (n = 99), non-systemic JIA (n = 169), infections (n = 51), other inflammatory diseases (n = 126), and acute lymphoblastic leukemia (ALL, n = 147). In addition, samples from 23 healthy controls were included.ResultsThe sCAL turbo assay correlated well with the MRP8/14 ELISA used in previous validation studies (r = 0.99, p < 0.001). It could reliably differentiate SJIA from all other diagnoses with significant accuracy (cutoff at 10,500 ng/ml, sensitivity 84%, specificity 94%, ROC area under curve 0.960, p < 0.001).ConclusionsSerum calprotectin analyses are a helpful tool supporting the diagnosis of SJIA in children with prolonged fever or inflammatory disease. Here, we show that an immuno-turbidimetric assay for detection of serum calprotectin on an automated laboratory instrument can be implemented in clinical laboratory settings to facilitate its use as a diagnostic routine test in clinical practice.

High-Dose Hydrocortisone Treatment Does Not Affect Serum C-Reactive Protein (CRP) Concentrations in Healthy Dogs.

Measuring C-reactive protein (CRP) in serum is a useful surrogate marker for assessing disease progression and treatment response in dogs with autoinflammatory diseases. Affected dogs often receive high-dose glucocorticoid treatment, but the effect of such treatment alone on serum CRP concentrations is unknown. We evaluated serum CRP concentrations via immunoassay (sandwich enzyme-linked immunosorbent assay and particle-enhanced turbidimetric immunoassay) in 12 healthy beagle dogs administered high-dose hydrocortisone (8 mg/kg q12 h) per os vs. placebo over 28 days (days 0, 1, 5, and 28) in a randomized parallel study design. Serum CRP concentrations slightly decreased during treatment or placebo but without a significant association with hydrocortisone administration (p = 0.761). Compared to baseline, serum CRP concentrations were decreased by >2.7-fold (minimum critical difference) in three hydrocortisone-treated dogs and two dogs in the placebo group on day 28, whereas an increase to >2.7-fold was seen in one dog receiving placebo. These results suggest a lack of confounding effects of high-dose hydrocortisone administration on serum CRP concentrations in healthy dogs. This might also hold in dogs with autoinflammatory conditions and/or administration of other high-dose corticosteroids, suggesting that CRP presents a suitable biomarker to monitor inflammatory disease processes. However, this needs confirmation by further studies evaluating corticosteroid-induced cellular (e.g., hepatic) transcriptome and proteome changes.

A-072 An All-Purpose Particle Enhanced Immunoturbidimetric Assay for the Comparable, Instrument-Independent Monitoring of Procalcitonin in the Management of Sepsis

Abstract Introduction Sepsis is a life-threatening organ dysfunction caused by a dysregulated host immune response to infection. It is a global health concern and a leading cause of death worldwide, affecting an estimate of 48.9 million people every year. Procalcitonin (PCT) revealed to be an indispensable parameter for the management of sepsis. However, recent analysis of EQA survey results suggested crucial deviations of PCT measurements in Germany and studies by the IFCC Working Group on standardization of PCT stated the need for a higher-order reference measurement procedure.Here we present the evaluation of the all-purpose particle enhanced immunoturbidimetric assay (PETIA) of DiaSys, which can be easily applied to every high-throughput clinical chemistry analyzer. The high diversity of applications and datasets match current diagnostic and regulative needs (IVDR). Procalcitonin FS enables the equivalent and instrument-independent monitoring of PCT in the management of sepsis. Methodology Evaluation of Procalcitonin FS was performed on seven clinical chemistry analyzer platforms (Jeol BioMajesty® JCA-BM6010/C, DiaSys respons®920 and respons®940, Roche cobas c501, Abbott Architect c8000 and Alinity c, Beckman Olympus AU680). Measurements were performed according to Clinical and Laboratory Standards Institute (CLSI) protocols. Results The onboard stability reaches 8 weeks (with chimney) on BioMajesty® JCA-BM6010/C, 20 days on respons®920 and respons®940, 21 weeks on cobas c501, 9 weeks on Architect, 12 weeks on Olympus AU680 and 13 weeks on Alinity c. The calibration stability is 4 weeks (with chimney) on BioMajesty® JCA-BM6010/C, 2 weeks (with chimney) on respons®920 and respons®940, 21 weeks on cobas c501, 3 weeks on Architect, 5 weeks on Olympus AU680 and 10 weeks on Alinity c. The intra-assay precision (CV%) ranges from ≤4.57% on respons®920 up to ≤7.19% on respons®940. The Total Precision (CV%) ranges from ≤3.99% on cobas c501 up to ≤9.62% on Olympus AU680. LOB ranges from 0.07 ng/mL on BioMajesty® JCA-BM6010/C up to 0.130 ng/mL on Olympus AU680. LOQ ranges from 0.20 ng/mL on Architect up to 0.277 ng/mL on Olympus AU680. The reagent displayed a linear recovery up to 50 ng/mL and no prozone effect was detected up to 1000 ng/mL on all seven analyzers. The Passing &amp; Bablok regression vs the reference test showed an excellent comparability/alignment between the methods (slope ≤1.11, intercept ≤0.156 ng/mL, R ≥ 0.986). Conclusion Evaluation work proves that DiaSys PETIA reagent Procalcitonin FS shows excellent performance on all seven clinical chemistry analyzers. In addition, the high comparability between the seven analyzers enables the reliable and instrument-independent monitoring of PCT in the management of sepsis, and thus offering a solution to the current landscape of deviating PCT measurement results.

Antibody-Conjugated Nanodiamonds as Dual-Functional Immunosensors for In Vitro Diagnostics.

Nanodiamonds (NDs) are carbon nanoparticles with a large refractive index, a high density, and exceptional chemical stability. When excited by green light, they can emit bright red fluorescence from implanted nitrogen-vacancy (NV) centers. Taking advantage of these properties, we have developed antibody-conjugated NDs as in vitro diagnostic sensors for two complementary assays: particle-enhanced turbidimetric immunoassay (PETIA) and spin-enhanced lateral flow immunoassay (SELFIA). To achieve this goal, monocrystalline diamond powders (∼100 nm in diameter) with or without NV implantation were first treated in molten KNO3 to reduce their size and shape inhomogeneity, followed by surface carboxylation in strong oxidative acids and non-covalent conjugation with antibodies in water. PETIA and SELFIA were carried out separately with a microplate reader and a magnetically modulated fluorescence analyzer. Using C-reactive protein (CRP) as the target antigen, we found that anti-CRP-conjugated NDs exhibited high colloidal stability over 1 month at 4 °C in buffer solution. The limits of detection for 3 μL of CRP sample solution were 0.06 μg/mL and 1 ng/mL with variation coefficients of less than 10 and 15% for PETIA and SELFIA, respectively. These two methods together provide a detection range of 1 ng/mL-10 μg/mL, potentially useful for clinical applications. This work represents the first practical use of rounded monocrystalline NDs as in vitro diagnostic reagents.

EQA: There's not a glitch in the matrix. Investigation of CRP bias on the Roche Cobas c701.

The Roche Cobas c Tina-quant C-Reactive Protein (CRP) IV method has shown persistent, significant negative bias compared to other methodologies in the UK NEQAS external quality assurance (EQA) scheme. The aim of this study was to compare CRP concentrations determined by this restandardized 4th generation assay with another commercially available method to assess the extent of any bias and impact on clinical cut-offs. CRP concentrations in 38 anonymized patient serum samples were determined by a particle-enhanced immunoturbidimetric assay on the Roche Cobas c701 module and a latex assay on the Beckman Coulter AU5800 analyser over a concentration range of 1-308mg/L. 97.4% samples analysed by the Roche Tina-quant CRP IV method demonstrated a negative bias compared to the Beckman AU latex method. The mean absolute bias was -4.12mg/L (range: -19.13-10.93mg/L; 95% CI: -17.25-9.01mg/L). The mean relative difference was -15.5% (range: -40-4%; 95% CI: -33.03-1.94%). This bias was seen across the range of samples assayed, including at clinically significant cut-offs of 5mg/L (-24% bias), 20mg/L (-5%) and 100mg/L (-13%). The negative bias of the Roche method demonstrated in the EQA scheme appears to reflect genuine differences in patient results, rather than an EQA matrix effect, despite re-standardization of the CRP assay. This report indicates that clinical cut-offs and reference ranges may be method-dependent, which should be reflected in published guidance and interpretation provided by laboratories.

Exploring Associations between C-Reactive Protein and Self-Reported Interoception in Major Depressive Disorder: A Bayesian Analysis.

Major depressive disorder (MDD) is associated with dysfunctional self-reported interoception (i.e., abnormal perception of the body's physiological state) and systemic inflammation, both of which adversely affect treatment response. In this study, we explored associations between C-reactive protein (CRP) and self-reported interoception, to gain more insight into the pathophysiology of interoceptive impairments in MDD. We also aimed to replicate previous findings on the associations of depression and fatigue severity with CRP. The study included 97 depressed individuals, who completed self-administered questionnaires (Multidimensional Assessment of Interoceptive Awareness (MAIA-2); Beck Depression Inventory-II, Multidimensional Fatigue Inventory). CRP concentrations were analyzed in the serum using a particle-enhanced turbidimetric immunoassay. We applied Bayesian inference to estimate robust effect parameters from posterior distributions based on MCMC sampling, and computed Bayes factors (BF10) as indices of relative evidence. The bivariate analysis supported evidence against associations between CRP and self-reported interoception (BF10 ≤ 0.32), except for one dimension (Not-Distracting: r = 0.11, BF10 > 0.43, absence of evidence). Positive correlations with overall depression (r = 0.21, BF10 = 3.19), physical fatigue (r = 0.28, BF10 = 20.64), and reduced activity (r = 0.22, BF10 = 4.67) were found. The multivariate analysis showed moderate evidence that low-grade inflammation predicted higher scores on the MAIA-2 Not-Worrying scale (β = 0.28, BF10 = 3.97), after controlling for relevant confounders. Inflammatory responses, as measured by CRP, may not be involved in the pathophysiology of dysfunctional self-reported interoception. However, systemic low-grade inflammation could potentially exert a protective effect against worries about pain or discomfort sensations. An immunological involvement in interoceptive impairments cannot be ruled out until future studies considering additional biomarkers of inflammation replicate our findings.

Bioinformatical Design and Performance Evaluation of a Nucleocapsid- and an RBD-Based Particle Enhanced Turbidimetric Immunoassay (PETIA) to Quantify the Wild Type and Variants of Concern-Derived Immunoreactivity of SARS-CoV-2.

Since SARS-CoV-2 emerged in December 2019 in Wuhan, the resulting pandemic has paralyzed the economic and cultural life of the world. Variants of concern (VOC) strongly increase pressure on public health systems. Rapid, easy-to-use, and cost-effective assays are essential to manage the pandemic. Here we present a bioinformatical approach for the fast and efficient design of two innovative serological Particle Enhanced Turbidimetric Immunoassays (PETIA) to quantify the SARS-CoV-2 immunoresponse. To confirm bioinformatical assumptions, an S-RBD- and a Nucleocapsid-based PETIA were produced. Sensitivity and specificity were compared for 95 patient samples using a BioMajesty™ fully automated analyzer. The S-RBD-based PETIA showed necessary specificity (98%) over the N protein-based PETIA (21%). Further, the reactivity and cross-reactivity of the RBD-based PETIA towards variant-derived antibodies of SARS-CoV-2 were assessed by a quenching inhibition test. The inhibition kinetics of the S-RBD variants Alpha, Beta, Delta, Gamma, Kappa, and Omicron were evaluated. In summary, we showed that specific and robust PETIA immunoassays can be rapidly designed and developed. The quantification of the SARS-CoV-2-related immunoresponse of variants (Alpha to Kappa) is possible using specific RBD assays. In contrast, Omicron revealed lower cross-reactivity (approx. 50%). To ensure the quantification of the Omicron variant, modified immunoassays appear to be necessary.

High levels of fecal calprotectin and C-reactive protein in patients with colitis.

Inflammatory bowel disease (IBD) with a poor prognosis may be due to persistent colitis. According to the latest guidelines, monitoring has become a part of the treatment process for colitis. Adequate monitoring of the patient's condition is necessary to determine the course of the disease to prevent the worsening of the condition and suppress the subclinical inflammatory process. This analytical study with a cross-sectional design was conducted to evaluate the activity of colitis using the results of C-reactive protein (CRP) and fecal calprotectin (FC) assays. FC levels were analyzed by ELISA, while CRP levels were analyzed using Siemens Flex particle-enhanced turbidimetric immunoassay. In 30 subjects with endoscopy and biopsy of colitis, 16 men and 14 women had a median age of 52.5 (18-70) years. The median FC value increased by 67 (7.3-722 g/g) and was positive (≥50 g/g) in 20 subjects (66.7%), and the mean CRP value was 13.64 mg/L, positive (10-15 mg/L) in 13 subjects (43.33%), and negative (<10 mg/L) in 17 subjects (56.67%). This study demonstrated that FC had a significant relationship with CRP (r=0.57; p<0.001) in patients with colitis. Assessing the levels of FC and CRP among patients with colitis can be useful to assess the worsening of symptoms early and reduce mortality and morbidity.

Using Residual Blood from the Arterial Blood Gas Test to Perform Therapeutic Drug Monitoring of Vancomycin: An Example of Good Clinical Practice Moving towards a Sustainable Intensive Care Unit.

Regarding sustainability in the intensive care unit (ICU), there is increasing interest in reducing material waste and avoiding unnecessary procedures. Therapeutic drug monitoring (TDM) of vancomycin, using a dedicated tube, is standard clinical care during treatment with vancomycin. Furthermore, in the ICU, on a daily basis, arterial blood gas (ABG) tests are frequently performed throughout the day. After analysis, a variable volume of blood is discarded. Lithium heparin (LiHep) syringes for ABG tests differ from normally used dipotassium ethylenediaminetetraacetic acid (K2EDTA) tubes. The primary objective was to compare both containers and validate the use of LiHep syringes. Secondary objectives were to evaluate the potential impact on saving materials, nursing time, and costs when implementing vancomycin TDM via LiHep syringes. Vancomycin analysis from sampling in lithium heparin (LiHep) syringes for ABG tests was validated and compared with the concentrations from conventional sampling in K2EDTA tubes. For method comparison, a Bland-Altman plot and Deming regression analysis were performed. The method was validated for inter- and intra-day precision and accuracy. Vancomycin was analyzed by means of the validated method using a particle-enhanced turbidimetric inhibition immunoassay (PETINIA) autoanalyzer. Furthermore, an analysis was conducted to evaluate the potential impact of implementing vancomycin sampling via ABG tests on savings in materials, nursing time, and costs. From 18 patients, 24 plasma samples in both K2EDTA tubes and LiHep syringes were obtained and compared. The mean relative difference between the two containers was -2.0% (-3.0 to -0.93%). Both the Deming regression analysis and the Bland-Altman plot met the acceptance criteria. Potentially, over 1000 blood draws and accompanying materials and packaging can be saved when vancomycin samples are obtained by means of scavenged LiHep syringes. The vancomycin analysis for LiHep syringes showed a total interday precision of 1.95% and an accuracy of 99.7%. The total intraday precision was 2.22%, and the accuracy was 99.2%. Accuracy and precision values were within the acceptance criteria of recovery 85 to 115% and ≤15%, respectively. No significant differences were found in vancomycin concentration between the two analyses, and the LiHep analysis was validated for further implementation in clinical care. Residual blood from ABG test samples can be used for TDM of vancomycin, resulting in a potential reduction of materials used and the number of blood draws. These results will contribute to a more sustainable TDM process with benefits for the patient.

Elevated Plasma D-Dimer Concentrations in Adults after an Outpatient-Treated COVID-19 Infection.

Elevated D-dimer plasma concentrations are common in hospitalized COVID-19 patients and are often associated with a worse prognosis, but it is not yet clear whether this also applies to outpatient cases. The present cross-sectional study evaluated D-dimer levels and their association with clinical parameters and inflammation biomarkers after a COVID-19 disease in individuals treated as outpatients. The study included 411 individuals (43.3% men) with an average age of 46.8 years (SD 15.2). Study participants who had acute COVID-19 disease at a median of 235 days (120; 323) ago were examined at the University Hospital Augsburg, Southern Germany, between 11/2020 and 05/2021. Plasma D-dimers were measured by a particle-enhanced immunoturbidimetric assay. Sixty-one subjects (15%) showed increased D-dimer concentrations (≥500 µg/L). Study participants with elevated D-dimer levels in comparison to subjects with levels in the reference range were significantly older, and more frequently reported a history of cardiovascular disease, hypertension, venous thromboembolism, and chronic venous insufficiency. In multivariable logistic regression analysis, CRP levels (OR 5.58 per mg/dL, 95% CI 1.77-17.60) and white blood cell count (OR 1.48 per nL, 95% CI 1.19-1.83) were significantly related to elevated D-dimers even after adjustment for multiple testing. However, acute or persistent symptoms were not significantly associated with increased D-dimers. Elevated D-dimer levels months after an acute COVID-19 disease seems to be associated with markers of inflammation. Further studies are needed to investigate the underlying pathophysiological mechanisms and consequences of prolonged D-dimer elevation in these patients.

Salivary Calprotectin Is not a Useful Biomarker to Monitor Disease Activity in Patients with Inflammatory Bowel Disease.

Non-invasive biomarkers are gaining interest for monitoring disease activity in patients with inflammatory bowel diseases (IBD). Fecal calprotectin is a reliable biomarker but patients often report the collection of feces being unpleasant and cumbersome. In this study, we aimed to assess if salivary calprotectin could be used as a non-invasive biomarker to determine disease activity instead of fecal calprotectin. In this cross-sectional explorative cohort study, stimulated saliva was collected from patients with an established IBD diagnosis and healthy controls. The concentration of calprotectin in saliva was determined by a particle-enhanced turbidimetric immunoassay. Intestinal disease activity was assessed with fecal calprotectin levels and the Harvey-Bradshaw Index (HBI) or Simple Clinical Colitis Activity Index (SCCAI). Missing data were handled using multiple imputation. Sixty-three patients (41 Crohn's disease and 22 ulcerative colitis) and 11 controls were included. Patients had a mean fecal calprotectin of 138.78 µg/g and a median salivary calprotectin of 1.87 mg/L. No significant correlation was found between salivary calprotectin and fecal calprotectin levels (p=0.495). When patients were stratified in two subgroups based on a fecal calprotectin cut-off value of 250 µg/g, there were no significant differences in salivary calprotectin levels between both patient groups (p=0.641) and between patients and healthy controls (p=0.248). Also, salivary, and fecal calprotectin levels were not significantly different when stratifying patients in two subgroups, active disease and remission, using HBI/SCCAI scores. Salivary calprotectin does not correlate to fecal calprotectin and disease activity scores in patients, making it unreliable for assessing IBD activity.