Novel open-source diagnostic tool offers affordable, reliable pathogen detection for resource-limited settings

A bottleneck in ensuring access to widespread molecular diagnostics, especially in low- and middle-income countries, has been the high cost and logistical complexities associated with rapid, point-of-care tests.

Now, a collaborative research effort outlined in a Life Science Alliance (LSA) study has addressed these challenges by developing a lyophilized (freeze-dried) open-source reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for pathogen detection. This method, successfully applied to COVID-19, aims to make the diagnostics more accessible and affordable globally.

In the study, “A lyophilized open-source RT-LAMP assay for molecular diagnostics in resource-limited settings,” scientists at the Vienna BioCenter in Vienna, Austria, and the West African Center for Cell Biology of Infectious Pathogens (WACCBIP) at the University of Ghana in Legon, Ghana, detail an RT-LAMP assay built entirely from nonproprietary enzymes including reverse transcriptase, DNA polymerase, and uracil DNA glycosylase.

The choice of ingredients was made to reduce the reliance on expensive commercial solutions that can be difficult to access in many parts of the world.

“By making reliable pathogen detection both affordable and independent of complex logistics like cold chains, our protocols empower laboratories and public health institutions worldwide to take control of their diagnostic capabilities, especially during outbreaks,” says co-first author Martin Matl, Ph.D. student at the Institute of Molecular Biotechnology (IMBA), Vienna BioCenter.

A remarkable feature of the new assay is its heat stability and robustness, even when stored at ambient or elevated temperatures. This bypasses the need for a cold chain during transport and storage, a major logistical and financial hurdle for deploying diagnostic tools in remote or resource-limited settings.

As a proof of concept, the lyophilized RT-LAMP reaction mixes were shipped to WACCBIP in Ghana, and demonstrated performance comparable to results obtained in Vienna, where the assay was developed.

The colorimetric assay provides a flexible and scalable point-of-care test that can be adapted for rapid detection of various pathogens. It also features a sample lysis/processing solution for direct testing that is on par with commercial formulations, and handles both swab and gargle samples with high sensitivity.

The open-source nature means the enzyme mixture for RT-LAMP can be self-produced, offering sensitivity comparable to commercially available kits. The system also incorporates measures to protect against contamination, a significant consideration for non-laboratory environments.

“To support real-world adoption, we envision several implementation models, ranging from centralized production and distribution of lyophilized reagents to decentralized in-lab preparation by regional facilities,” says co-corresponding author Gordon A. Awandare, Director of WACCBIP at the University of Ghana.

“By validating the system’s performance across diverse sample types and international settings, we demonstrate that the platform is both robust and suitable for field implementation.”

The team has deposited E. coli expression vectors for the His-tagged versions of the three wild-type enzymes with Addgene and provides standard protocols for protein expression and purification on their website to further facilitate widespread adoption.

A bottleneck in ensuring access to widespread molecular diagnostics, especially in low- and middle-income countries, has been the high cost and logistical complexities associated with rapid, point-of-care tests.

Now, a collaborative research effort outlined in a Life Science Alliance (LSA) study has addressed these challenges by developing a lyophilized (freeze-dried) open-source reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for pathogen detection. This method, successfully applied to COVID-19, aims to make the diagnostics more accessible and affordable globally.

In the study, “A lyophilized open-source RT-LAMP assay for molecular diagnostics in resource-limited settings,” scientists at the Vienna BioCenter in Vienna, Austria, and the West African Center for Cell Biology of Infectious Pathogens (WACCBIP) at the University of Ghana in Legon, Ghana, detail an RT-LAMP assay built entirely from nonproprietary enzymes including reverse transcriptase, DNA polymerase, and uracil DNA glycosylase.

The choice of ingredients was made to reduce the reliance on expensive commercial solutions that can be difficult to access in many parts of the world.

“By making reliable pathogen detection both affordable and independent of complex logistics like cold chains, our protocols empower laboratories and public health institutions worldwide to take control of their diagnostic capabilities, especially during outbreaks,” says co-first author Martin Matl, Ph.D. student at the Institute of Molecular Biotechnology (IMBA), Vienna BioCenter.

A remarkable feature of the new assay is its heat stability and robustness, even when stored at ambient or elevated temperatures. This bypasses the need for a cold chain during transport and storage, a major logistical and financial hurdle for deploying diagnostic tools in remote or resource-limited settings.

As a proof of concept, the lyophilized RT-LAMP reaction mixes were shipped to WACCBIP in Ghana, and demonstrated performance comparable to results obtained in Vienna, where the assay was developed.

The colorimetric assay provides a flexible and scalable point-of-care test that can be adapted for rapid detection of various pathogens. It also features a sample lysis/processing solution for direct testing that is on par with commercial formulations, and handles both swab and gargle samples with high sensitivity.

The open-source nature means the enzyme mixture for RT-LAMP can be self-produced, offering sensitivity comparable to commercially available kits. The system also incorporates measures to protect against contamination, a significant consideration for non-laboratory environments.

“To support real-world adoption, we envision several implementation models, ranging from centralized production and distribution of lyophilized reagents to decentralized in-lab preparation by regional facilities,” says co-corresponding author Gordon A. Awandare, Director of WACCBIP at the University of Ghana.

“By validating the system’s performance across diverse sample types and international settings, we demonstrate that the platform is both robust and suitable for field implementation.”

The team has deposited E. coli expression vectors for the His-tagged versions of the three wild-type enzymes with Addgene and provides standard protocols for protein expression and purification on their website to further facilitate widespread adoption.