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Sickle cell disease (SCD) is a group of common, life-threatening disorders caused by a point mutation in the β globin gene. Early diagnosis through newborn and early childhood screening, parental education, and preventive treatments are known to reduce mortality. However, the cost and complexity of conventional diagnostic methods limit the feasibility of early diagnosis for SCD in resource-limited areas worldwide. Although several point-of-care tests are commercially available, most are antibody-based tests, which cannot be used in patients who have recently received a blood transfusion. Here, we describe the development of a rapid, low-cost nucleic acid test that uses real-time fluorescence to detect the point mutation encoding hemoglobin S (HbS) in one round of isothermal recombinase polymerase amplification (RPA). When tested with a set of clinical samples from SCD patients and healthy volunteers, our assay demonstrated 100% sensitivity for both the βA globin and βS globin alleles and 94.7 and 97.1% specificities for the βA globin allele and βS globin allele, respectively (n = 91). Finally, we demonstrate proof-of-concept sample-to-answer genotyping of genomic DNA from capillary blood using an alkaline lysis procedure and direct input of diluted lysate into RPA. The workflow is performed in

INTRODUCTION: Cervical cancer mortality rates remain high in low- and middle-income countries (LMICs) and other medically underserved areas due to challenges with implementation and sustainability of routine screening, accurate diagnosis, and early treatment of preinvasive lesions. AREAS COVERED: In this review, we first discuss the standard of care for cervical cancer screening and diagnosis in high- and low-resource settings, biomarkers that correlate to cervical precancer and cancer, and needs for new tests. We review technologies for screening and diagnosis with a focus on tests that are already in use in LMICs or have the potential to be adapted for use in LMICs. Finally, we provide perspectives on the next five years of technology development for improved cervical cancer screening and diagnosis in LMICs. EXPERT OPINION: Innovation toward improved molecular and imaging tests is needed to enable effective, affordable see-and-treat approaches to detect and treat cervical precancer in a single visit. Current molecular tests remain too complex and/or costly for widespread use. Especially with imaging tests, decision support may improve performance of new technologies.

Traditional methods to detect and quantify nucleic acids rely on polymerase chain reaction (PCR) and require the use of expensive thermocyclers with integrated fluorescence detection of amplicons. Isothermal nucleic acid amplification technologies eliminate the need for thermal cycling; however, fluorescence-based detection of products is still required for real-time, quantitative results. Several portable isothermal heaters with integrated fluorescence detection are now commercially available; however, the cost of these devices remains a significant barrier to widespread adoption in resource-limited settings. Described here is a protocol for the design and assembly of a modular, low-cost fluorimeter constructed from off-the-shelf components. Enclosed in a compact 3D printed housing, the fluorimeter is designed to be placed atop a commercially available heat block holding a PCR tube. The fluorimeter described here was optimized to detect fluorescein isothiocyanate (FITC) dye, but the system can be modified for use with dyes commonly used as reporters in real-time nucleic acid amplification reactions. Clinical applicability of the system is demonstrated by performing real-time nucleic acid detection with two isothermal amplification technologies: recombinase polymerase amplification (RPA) for detection of positive control DNA provided in a commercial kit and reverse transcription loop-mediated isothermal amplification (RT-LAMP) for detection of clinically meaningful levels of SARS-CoV-2 RNA.

Each day, approximately 830 women and 7,400 newborns die from complications during pregnancy and childbirth. Improving maternal and neonatal health will require bringing rapid diagnosis and treatment to the point of care in low-resource settings. However, to date there are few diagnostic tools available that can be used at the point of care to detect the leading causes of maternal and neonatal mortality in low-resource settings. Here we review both commercially available diagnostics and technologies that are currently in development to detect the leading causes of maternal and neonatal mortality, highlighting key gaps in development where innovative design could increase access to technology and enable rapid diagnosis at the bedside.

Background Sickle cell anaemia is a common, life-threatening disorder caused by a point mutation in the β globin gene. The high cost and complexity of conventional diagnostic methods limit the scope and sustainability of newborn screening for sickle cell anaemia and other haemoglobin disorders in resource-limited areas. Although several point-of-care tests are commercially available, most are antibody-based tests, which cannot be used in patients who have recently received a blood transfusion. Additionally, very low levels of sickle haemoglobin (HbS) in premature babies and some newborns may cause errors in immunoassays. DNA analysis provides the most reliable approach to diagnose haemoglobin disorders. However, DNA analysis is not currently available at the point of care due to its cost and complexity. Here, we describe a rapid, low-cost nucleic acid test that uses real-time fluorescence to detect the point mutation encoding HbS in one round of isothermal recombinase polymerase amplification. Methods Venous and capillary blood samples were obtained from healthy volunteers who were recruited by word of mouth. Venous blood samples from patients with varying genotypes seen in the Sickle Cell Disease Program at Texas Children's Hematology Center were collected and deidentified under IRB-approved exemption. Genotypes, as determined by qualitative isoelectric focusing combined with quantitative high performance liquid chromatography (HPLC) to confirm a diagnosis, were provided with the blood samples. The assay was evaluated with genomic DNA extracted from a set of blood samples of varying genotypes using the Axxin T8-ISO, a low-cost, field-deployable benchtop fluorimeter. Findings The developed assay demonstrated 100% sensitivity for both the βA globin and βS globin alleles, and 94·7% and 97·1% specificity for the βA globin allele and βS globin allele, respectively (n=91). We also demonstrate a complete sample-to-answer workflow using a two-step lysis method and direct input of lysate into the allele-specific assay, and genotyping of blood samples in an open-source, custom built fluorimeter whose components cost less than US$850. Interpretation This sample-to-answer nucleic acid test has the potential to be adapted to other disease-causing point mutations in genomic DNA. The technology described could streamline diagnosis in resource-limited settings of patients with sickle cell anaemia who have recently undergone a blood transfusion, with a comparable cost to protein-based tests. Funding NIH K23-HL148548-01A1.

Background: The global COVID-19 pandemic caused by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has highlighted the need for rapid, accurate, and accessible diagnostics to enable timely treatment and outbreak control. However, current diagnostic tests based on RT-qPCR are insufficient to meet the global testing demand because of their high cost and complexity and supply chain shortages. Reverse transcription loop-mediated isothermal amplification (RT-LAMP) is a promising alternative to RT-qPCR because of its sensitivity, speed, and robustness to sample inhibitors. Here, we describe the development and optimisation of a sample-to-answer workflow, including a simple lysis and inactivation protocol that provides results in

Background The detection and quantification of nucleic acids traditionally relies on PCR, which requires the use of expensive thermocyclers with integrated fluorescence detection of amplicons. While isothermal nucleic acid amplification technologies eliminate the need for thermocycling, fluorescence-based detection of products is still required for real-time, quantitative readout. Several commercially available isothermal heaters with integrated fluorescence detection exist, but the cost of these devices remains a significant barrier to the use of isothermal nucleic acid amplification in low-resource settings. A low-cost device for nucleic acid amplification detection could be used to contain the spread of the SARS-CoV-2 virus during the current pandemic as well as other disease targets in the future. Increasing COVID-19 testing capacity is particularly relevant when variations of the virus that appear to be more infectious are spreading rapidly and efforts to scale up delivery of COVID-19 vaccines are in the early stages. Here, we describe a modular, low-cost fluorimeter for use in resource-limited settings that lack access to and infrastructure for traditional testing platforms. Methods We present a fluorimeter that is constructed from off-the-shelf components enclosed in a compact 3D printed housing that removes the need for specialised optical alignment. The fluorimeter is designed to be placed atop a commercially available heat block holding a PCR-tube and was optimised to detect fluorescein (FITC) dye. The modular design allows it to be easily adapted for use with other dyes commonly used as reporters in nucleic acid amplification reactions. Findings Clinical applicability of the system was demonstrated using the system to measure amplification with two different isothermal amplification technologies with significantly different temperature, volume, and fluorescence requirements: recombinase polymerase amplification (RPA) and reverse transcription loop-mediated isothermal amplification (RT-LAMP). RPA detection was demonstrated with a positive control DNA provided in a commercial kit while RT-LAMP detection of clinically meaningful levels of SARS-CoV-2 RNA was demonstrated in a custom RT-LAMP assay. In a preliminary analysis of nasopharyngeal swab samples from 74 individuals, the custom RT-LAMP assay showed a positive agreement of 92·3% and a negative agreement of 91·4% with the SARS-CoV-2 RT-qPCR test developed by the US Centers for Disease Control and Prevention. Interpretation This open-source system was designed to improve accessibility to nucleic acid testing with isothermal amplification methods. The system can be easily produced using only a low-end 3D printer. Moreover, the use of single-board computers obviates the need to produce printed circuit boards. At a time when global supply chains are at their most fragile, open-source diagnostic equipment has the potential to reduce pandemic-related health inequities. Funding International AIDS vaccine Initiative IAVI and USAID: award AID-OAA-A16-00032.

There is a need to develop inexpensive, portable and easy-to-use devices for viral sample processing for resource-limited settings. Here we offer a solution to efficient virus capture by incorporating macroporous materials with regular structures into microfluidic devices for affinity chromatography. Two-dimensional simulations were first conducted to investigate the effects of two structures, a nanopost array and a spherical pore network, on nanoparticle capture. Then, the two structures were created in polymers by templating anodic aluminum oxide films and 3D close-packed silica particles, respectively. When the microdevices containing functionalized porous materials were tested for human immunodeficiency virus (HIV) isolation, capture efficiencies of 80–99% were achieved under a continuous flow. Comparatively, functionalized flatbed microchannels captured around 10% of HIV particles. As the characteristic dimensions of the nanostructures are tunable, such devices can be adapted for the capture of different submicron bioparticles. The high capture efficiency and easy-to-operate nature suit the needs of resource-limited settings and may find applications in point-of-care diagnostics.