A definitive diagnosis of Alzheimer’s disease (AD), even in the presence of co-morbid neuropathology (occurring in > 50% of AD cases), is a significant unmet medical need that has obstructed the discovery of effective AD therapeutics. An AD-biomarker, the Morphometric Imaging (MI) assay on cultured skin fibroblasts, was used in a double-blind, allcomers (ages 55–90) trial of 3 patient cohorts: AD dementia patients, N = 25, all autopsy confirmed, non-AD dementia patients, N = 21—all autopsy or genetically confirmed; and non-demented control (AHC) patients N = 27. Fibroblasts cells isolated from 3-mm skin punch biopsies were cultured on a 3-D Matrigel matrix with movement dynamics quantified by image analysis. From counts of all aggregates (N) in a pre-defined field image and measures of the average area (A) of aggregates per image, the number-to-area ratios in a natural logarithmic form Ln(A/N) were determined for all patient samples. AD cell lines formed fewer large aggregates (cells clustered together) than non-AD or AHC cell lines. The cut-off value of Ln(A/N) = 6.98 was determined from the biomarker values of non-demented apparently healthy control (AHC) cases. Unequivocal validation by autopsy, genetics, and/or dementia criteria was possible for all 73 patient samples. The samples were collected from multiple centers—four US centers and one center in Japan. The study found no effect of center-to-center variation in fibroblast isolation, cell growth, or cell aggregation values (Ln(A/N)). The autopsy-confirmed MI Biomarker distinguished AD from non-AD dementia (non-ADD) patients and correctly diagnosed AD even in the presence of other co-morbid pathologies at autopsy (True Positive = 25, False Negative = 0, False Positive = 0, True Negative = 21, and Accuracy = 100%. Sensitivity and specificity were calculated as 100% (95% CI = 84 to 100.00%). From these findings, the MI assay appears to detect AD with great accuracy—even with abundant co-morbidity.

Lipoarabinomannan (LAM), a component of the Mycobacterium tuberculosis (MTB) cell wall, is detectable in the urine of MTB infected patients with active tuberculosis (TB). LAM-specific antibodies (Igs) have been developed by a variety of traditional and recombinant methods for potential use in a rapid diagnostic test (RDT). We evaluated the analytical performance of the TB LAM Igs to identify pairs that offer superior performance over existing urine LAM tests. We assessed 25 new and 4 existing Igs in a matrixed format using a multiplex electrochemiluminescence-based liquid immunoassay. A total of 841 paired Ig combinations were challenged with in vitro cultured LAM (cLAM) derived from MTB strains representing diverse phylogenetic lineages, alongside urinary LAM (uLAM) from the urine of adults with active pulmonary TB. Analytical sensitivity of down-selected Ig pairs was determined using MTB Aoyama-B cLAM, while diagnostic accuracy was determined using clinical samples. When testing cLAM, the reactivity of Ig pairs was similar across MTB lineages 1–4 but lineage 5:6 had significantly more reactivity among Ig pairs. Overall, 41 Ig pairs had a strong binding affinity to cLAM, as compared to the reference pair of S4-20/A194-01, and 28 Ig pairs therein exhibited a strong affinity for both cLAM and uLAM. Retrospective testing on clinical urine specimens demonstrated varying sensitivities (12–80%) and specificities (14–100%). The five top pairs had a similar analytical limit of detection to the reference pair but in four instances, the sensitivity and specificity with clinical uLAM samples was poor. Overall, epitopes presented by uLAM are different from cLAM, which may affect antibody performance when testing uLAM in patient samples. Several new Ig pairs had similar ranges of high sensitivity to cLAM but overall, there were no new candidate Ig pairs identified in this round of screening with increased performance with uLAM as compared to an existing optimal pair.

The growth of resistance to multiple herbicides in grass weeds is a major threat to global cereal production and in the UK, is epitomized by the loss of control of blackgrass (Alopecurus myosuroides), causing losses in winter wheat production equating to 5% of national consumption. With an urgent need to develop new black-grass management tools, we have developed a lateral flow assay (LFA) that can predict resistance to multiple herbicides within 10 min.

AbstractLipoarabinomannan (LAM), a component of the Mycobacterium tuberculosis (MTB) cell wall, is detectable in the urine of MTB infected patients with active tuberculosis (TB). LAM-specific antibodies (Igs) have been developed by a variety of traditional and recombinant methods for potential use in a rapid diagnostic test (RDT). We evaluated the analytical performance of the TB LAM Igs to identify pairs that offer superior performance over existing urine LAM tests. We assessed 25 new and 4 existing Igs in a matrixed format using a multiplex electrochemiluminescence-based liquid immunoassay. A total of 841 paired Ig combinations were challenged with in vitro cultured LAM (cLAM) derived from MTB strains representing diverse phylogenetic lineages, alongside urinary LAM (uLAM) from the urine of adults with active pulmonary TB. Analytical sensitivity of down-selected Ig pairs was determined using MTB Aoyama-B cLAM, while diagnostic accuracy was determined using clinical samples. When testing cLAM, the reactivity of Ig pairs was similar across MTB lineages 1-4 but lineage 5:6 had significantly more reactivity among Ig pairs. Overall, 41 Ig pairs had a strong binding affinity to cLAM, as compared to the reference pair of S4-20/A194-01, and 28 Ig pairs therein exhibited a strong affinity for both cLAM and uLAM. Retrospective testing on clinical urine specimens demonstrated varying sensitivities (12-80%) and specificities (14-100%). The five top pairs had a similar analytical limit of detection to the reference pair but in four instances, the sensitivity and specificity with clinical uLAM samples was poor. Overall, epitopes presented by uLAM are different from cLAM, which may affect antibody performance when testing uLAM in patient samples. Several new Ig pairs had similar ranges of high sensitivity to cLAM but overall, there were no new candidate Ig pairs identified in this round of screening with increased performance with uLAM as compared to an existing optimal pair.

AbstractWith the distribution of COVID-19 vaccinations across the globe and the limited access in many countries, quick determination of an individual’s antibody status could be beneficial in allocating limited vaccine doses in low- and middle-income countries (LMIC). Antibody lateral flow tests (LFTs) have potential to address this need as a quick, point of care test, they also have a use case for identifying sero-negative individuals for novel therapeutics, and for epidemiology. Here we present a proof-of-concept evaluation of eight LFT brands using sera from 95 vaccinated individuals to determine sensitivity for detecting vaccination generated antibodies. All 95 (100%) participants tested positive for anti-spike antibodies by the chemiluminescent microparticle immunoassay (CMIA) reference standard post-dose two of their SARS-CoV-2 vaccine: BNT162b2 (Pfizer/BioNTech, n=60), AZD1222 (AstraZeneca, n=31), mRNA-1273 (Moderna, n=2) and Undeclared Vaccine Brand (n=2). Sensitivity increased from dose one to dose two in six out of eight LFTs with three tests achieving 100% sensitivity at dose two in detecting anti-spike antibodies. These tests are quick, low-cost point-of-care tools that can be used without prior training to establish antibody status and may prove valuable for allocating limited vaccine doses in LMICs to ensure those in at risk groups access the protection they need. Further investigation into their performance in vaccinated peoples is required before more widespread utilisation is considered.

ABSTRACTBackground: The emergence of variants of concern (VOCs) requires an ongoing assessment of the performance of antigen lateral flow tests (Ag-RDTs). The limit of detection (LOD) of 32 Ag-RDTs was evaluated using the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Gamma variant. Methods: Ag-RDTs were performed according to the manufacturer’s instructions with a clinical isolate of the Gamma variant. Results: Twenty-eight of the 32 Ag-RDTs exceeded the World Health Organization criteria. Conclusions: This comprehensive analytical evaluation of Ag-RDTs demonstrated that the test performance was maintained with Gamma VOC.

This study evaluated the validity and utility of antigen-detection rapid diagnostic tests (Ag-RDTs) for SARS-CoV-2 in elite sports. The data on utility, ease of use and application for Ag-RDTs as a new testing format were positive from players and staff. This evaluation was limited by the low prevalence of SARS-CoV-2 circulating within the three squads. This study highlights the need for continued service evaluations for SARS-CoV-2 Ag-RDTs in elite sport settings.

AbstractThe limit of detection (LOD) of thirty-two antigen lateral flow tests (Ag-RDT) were evaluated with the SARS-CoV-2 Gamma variant. Twenty-eight of thirty-two Ag-RDTs exceeded the World Health Organization criteria of an LOD of 1.0×106 genome copy numbers/ml and performance was equivalent as with the 2020 B.1 lineage and Alpha variant.

As the second year of the COVID-19 pandemic begins, it remains clear that a massive increase in the ability to test for SARS-CoV-2 infections in a myriad of settings is critical to controlling the pandemic and to preparing for future outbreaks. The current gold standard for molecular diagnostics is the polymerase chain reaction (PCR), but the extraordinary and unmet demand for testing in a variety of environments means that both complementary and supplementary testing solutions are still needed. This review highlights the role that loop-mediated isothermal amplification (LAMP) has had in filling this global testing need, providing a faster and easier means of testing, and what it can do for future applications, pathogens, and the preparation for future outbreaks. This review describes the current state of the art for research of LAMP-based SARS-CoV-2 testing, as well as its implications for other pathogens and testing. The authors represent the global LAMP (gLAMP) Consortium, an international research collective, which has regularly met to share their experiences on LAMP deployment and best practices; sections are devoted to all aspects of LAMP testing, including preanalytic sample processing, target amplification, and amplicon detection, then the hardware and software required for deployment are discussed, and finally, a summary of the current regulatory landscape is provided. Included as well are a series of first-person accounts of LAMP method development and deployment. The final discussion section provides the reader with a distillation of the most validated testing methods and their paths to implementation. This review also aims to provide practical information and insight for a range of audiences: for a research audience, to help accelerate research through sharing of best practices; for an implementation audience, to help get testing up and running quickly; and for a public health, clinical, and policy audience, to help convey the breadth of the effect that LAMP methods have to offer.

This review article concerns the production of recombinant antibody fragments for applications mainly in the diagnostic sector. The so-called “point of care diagnostics” is very important for timely diagnosis and treatment, thus being able to save lives and resources. There is intense pressure for more accurate and less expensive rapid diagnostic tests, with a value preferably <$1. Thus, the large-scale cost-effective production of recombinant antibodies is vital. The importance of Escherichia coli toward the production of inexpensive rapid tests will be explained in this review paper. Details about the different strains of E. coli, the strategies used for the insertion and the expression of recombinant proteins, and the challenges that still exist are provided. Afterward, the importance of the expression scale and culture parameters in the final yield of the antibodies are examined. From this analysis, it appears that for good yields of recombinant antibodies, aside from appropriate gene transfer and expression, the culturing parameters are of paramount importance. Larger scale production is more favorable, mainly due to the higher cell densities that can be achieved. Yields of functional Fab fragments in the range of 10–20 mg/L are considered good in shake flasks, whereas in bioreactors can be up to 1–2 g/L. An amount of 10–500 mg of such antibody per million rapid tests is required. Despite the substantial importance of the production of the antibodies and their fragments, their downstream processing should be appropriately considered from the beginning for achieving the target value of the final rapid diagnostic tests.