Project Summary/Abstract: An antigen test for Neisseria gonorrhoeae (Ng) that is accurate, simple, scalable, non-invasive, rapid, and effective at the point-of-care (POC), would transform public health decisions. Although antigen detection rapid diagnostic tests (Ag-RDT's) for Ng are available, they have poor sensitivity (50% or less) compared to gold standard nucleic acid amplification tests (NAAT). A NAAT based POC test was recently cleared by the US FDA, however its use in resource-poor settings is limited by its high cost. To overcome these barriers for the first time, we will harness ultrabright fluorescent nanoconstructs, called plasmonic-fluors, a recent breakthrough from our labs that enables such antigen testing with sensitivity that matches NAATs. The objective of this proposal is to develop an ultrasensitive plasmon-enhanced lateral flow assay (p-LFA) that detects the lipoprotein H.8 antigen present on Ng surface. The rationale underlying this proposal is that a highly sensitive Ag-RDT that can match the accuracy of NAAT could meet the critical need for diagnosing Ng infection at the POC and in resource-limited settings. We will achieve this by pursuing two specific aims: 1) Develop and optimize the p-LFA for detecting Ng lipoprotein H.8 antigen in urine samples obtained from healthy people. 2) Determine the diagnostic accuracy of the Ng antigen test compared to gold standard NAAT in urine specimens from patients with suspected Ng infection. The proposed approach is innovative in that it harnesses plasmonic-fluor as a fluorescent nanolabel in a LFA as opposed to the conventional gold nanoparticles, which provide only a weak colorimetric signal. With over 1000-fold improvement in sensitivity, this approach has the potential to transform antigen testing. The proposed research is significant as the novel p-LFA will meet the critical needs for widespread Ng testing scalability, speed, and low cost. The expected outcome of this work is a simple, non-invasive, Ag- RDT for Ng infection, implemented as a LFA that can replace NAAT by virtue of its accuracy and low cost in resource-limited settings. This test will have a tremendous positive impact immediately as it will be applicable to large scale Ng testing with an inexpensive test strip and minimal battery-powered portable equipment, without relying on skilled personnel. Furthermore, this test will serve as a prototype for ultrasensitive antigen tests applicable to many other infectious diseases in addition to the possibility of incorporating multiple antigens on a LFA for diagnosing other sexually transmitted infections such as chlamydia and trichomoniasis in addition to Ng.