A simple and inexpensive fluorescence-based technique has been developed for high-throughput antimalarial drug screening. This method uses SYBR Green I, a dye that enhances fluorescence upon contact with Plasmodium DNA, to measure parasite growth. The study compared this fluorescence assay with a standard radioisotopic method using known antimalarial agents against the Plasmodium falciparum strain D6. Both methods were used to determine the effective concentration of drug that resulted in a 50% reduction in observed counts (EC50) after 48 hours of parasite growth. The EC50 values for various drugs were similar or identical using both techniques, suggesting the fluorescence assay is a viable alternative for high-throughput screening. The fluorescence assay is simple, cost-effective, and robust, making it suitable for automated analysis. It relies on the contrast between host erythrocytes (lacking DNA and RNA) and malaria parasites (which have nucleic acids and are stained by the dye). The assay involves a one-step process, eliminating the need for complex protocols or expensive equipment. The study demonstrated that the fluorescence method is reliable, with low variability between triplicate wells and consistent EC50 values even after a freeze-thaw cycle. The method is also linear with parasitemia levels, showing a strong correlation between fluorescence units and parasite levels. The fluorescence-based method, called the SYBR Green I-based fluorescence (MSF) assay, offers advantages over radioisotopic methods, including lower cost, simplicity, and reduced need for specialized equipment. It has been successfully used in the laboratory to replace radioisotopic assays and to perform numerous high-throughput tests. The method is adaptable to various conditions, including different parasite strains and culture conditions. While some drugs may interfere with fluorescence, the overall results are consistent with radioisotopic methods, indicating the assay's reliability. The study highlights the potential of the fluorescence-based method as a practical and effective tool for antimalarial drug screening, particularly in resource-limited settings. It provides a cost-effective and efficient alternative to traditional radioisotopic methods, making it suitable for high-throughput screening and field applications. The method is also safe, with SYBR Green I being less mutagenic than ethidium bromide, and proper disposal protocols are recommended to ensure safety. The findings suggest that the fluorescence-based assay is a promising approach for antimalarial drug screening, with the potential to be widely applicable in both research and clinical settings.A simple and inexpensive fluorescence-based technique has been developed for high-throughput antimalarial drug screening. This method uses SYBR Green I, a dye that enhances fluorescence upon contact with Plasmodium DNA, to measure parasite growth. The study compared this fluorescence assay with a standard radioisotopic method using known antimalarial agents against the Plasmodium falciparum strain D6. Both methods were used to determine the effective concentration of drug that resulted in a 50% reduction in observed counts (EC50) after 48 hours of parasite growth. The EC50 values for various drugs were similar or identical using both techniques, suggesting the fluorescence assay is a viable alternative for high-throughput screening. The fluorescence assay is simple, cost-effective, and robust, making it suitable for automated analysis. It relies on the contrast between host erythrocytes (lacking DNA and RNA) and malaria parasites (which have nucleic acids and are stained by the dye). The assay involves a one-step process, eliminating the need for complex protocols or expensive equipment. The study demonstrated that the fluorescence method is reliable, with low variability between triplicate wells and consistent EC50 values even after a freeze-thaw cycle. The method is also linear with parasitemia levels, showing a strong correlation between fluorescence units and parasite levels. The fluorescence-based method, called the SYBR Green I-based fluorescence (MSF) assay, offers advantages over radioisotopic methods, including lower cost, simplicity, and reduced need for specialized equipment. It has been successfully used in the laboratory to replace radioisotopic assays and to perform numerous high-throughput tests. The method is adaptable to various conditions, including different parasite strains and culture conditions. While some drugs may interfere with fluorescence, the overall results are consistent with radioisotopic methods, indicating the assay's reliability. The study highlights the potential of the fluorescence-based method as a practical and effective tool for antimalarial drug screening, particularly in resource-limited settings. It provides a cost-effective and efficient alternative to traditional radioisotopic methods, making it suitable for high-throughput screening and field applications. The method is also safe, with SYBR Green I being less mutagenic than ethidium bromide, and proper disposal protocols are recommended to ensure safety. The findings suggest that the fluorescence-based assay is a promising approach for antimalarial drug screening, with the potential to be widely applicable in both research and clinical settings.