In this technical assessment of a novel GFAP LFA in a cohort including samples from TBI patients and healthy volunteers, the Upfront Dx LVOne GFAP LFA demonstrated a high PPV (90%, 95% CI: 77%, 87%) and sensitivity (95%, 95% CI: 83%, 99%) in the detection of samples with a GFAP concentration exceeding the manufacturer’s reported lower limit of detection (0.2 ng/ml). Furthermore, there was a significant positive association between GFAP concentrations as measured using the Quanterix Simoa® Human Neurology 4-Plex B assay and the semiquantitative score provided by the LVOne GFAP LFT.

In a prior study, a different iteration of the LVOne GFAP LFA, employing a marginally higher threshold of GFAP (0.213 ng/ml) to indicate a positive test result, was used to assess plasma GFAP concentration in patients with suspected stroke. In this study, a significant positive correlation was observed between Abbott iSTAT TBI GFAP quantifications and the LVOne GFAP LFA qualitative results (n = 20, Rho = 0.86) with a single false positive and no false negatives (Gaude et al. 2025). These findings are largely consistent with the analysis presented in this study, which has extended the assessment of the LVOne GFAP LFA to a different disease state, used serum instead of plasma samples and compared it to a different lab-based GFAP assay. Overall, we observed a similar positive correlation between the semiquantitative LVOne GFAP test result and the lab quantified serum GFAP concentration. We did, however, observe a greater proportion of both false positives and false negatives than seen previously with the LVOne GFAP LFA. Of the false positives observed in our study, one was a TBI patient with an acute subdural haemorrahge on CT and three were healthy controls. The higher proportion of false positives and negatives in our analysis, in comparison to the prior study of the LVOne, may be due to the use of different iterations of the LVOne with marginally different GFAP thresholds and different test line intensities to indicate a positive LFT result.

Although they represented less than 5% of the overall sample, the false negatives identified in our analysis are of particular concern. Whilst multiple stages of further research and refinement are required before clinical use of the LVOne in a TBI population, it remains noteworthy that both patients with false negative results had intracranial injuries requiring neurosurgical intervention. If the LVOne LFT had been used to triage these patients, such results could have led to inappropriate conveyance and delays in care. Future research should incorporate clinical perspectives to define an acceptable threshold for false negatives and to establish what proportion of missed injuries or incorrect transfers can be tolerated. Whilst the answer to this will depend on the specific clinical application of the device, it remains a key question for future investigation.

Although prior, and ongoing, study of the LVOne GFAP LFT has focussed on LVO stroke, GFAP is a biomarker of considerable promise in TBI. Several large observational studies have found GFAP to be sensitive for the detection of traumatic pathology on brain CT (Bazarian et al. 2018; Czeiter et al. 2020), associate with the burden of parenchymal disease (Whitehouse et al. 2022), and be sensitive for the detection of CT occult structural damage later seen on acute MRI (Yue et al. 2019). Furthermore, of particular interest in relation to the potential clinical uses of a GFAP LFT, a hyperacute rise in GFAP has been demonstrated in a prehospital study where samples were collected within 30 min of sustaining a suspected moderate-to-severe TBI, with predictive ability demonstrated for the presence of CT pathology and the likelihood of requiring acute neurosurgical intervention (Papa et al. 2024).

It should be noted that the sampling performed in this study was for serum samples, which required laboratory preparation. Separate validation of the LVOne GFAP LFA will be necessary for the use of whole blood and/or capillary samples. However, although there are commercially available PoC platforms offering quantitative GFAP analysis of whole blood in a similar time-frame as the LVOne GFAP LFT (Kobeissy et al. 2024), should the LVOne, or similar tests, be validated for use with whole blood or capillary samples they may offer a distinct set of advantages which may complement or expand current testing options. Notably, LFTs do not require a dedicated analyser or cartridge-based system, which could reduce overall cost, minimise training requirements, and eliminate the need for ongoing calibration or maintenance. These features may enhance the device’s usability in resource-limited or austere clinical environments, such as pre-hospital settings, sports sidelines or low- and middle-income countries (LMICs), where infrastructure for conventional testing platforms may be lacking.

In the pre-hospital context, biomarker testing could facilitate more appropriate triage decisions, such as directing patients to neurosurgical versus non-neurosurgical centres, and allow for early activation of specialist teams, including neurosurgery, prior to arrival. (Tepas et al. 2013) It is therefore relevant that the LVOne test is currently undergoing prospective validation of capillary testing for GFAP and D-Dimer for the triaging of ischaemic stroke due to large vessel occlusion (ISRCTN12414986). Similarly, in LMICs where access to CT imaging is limited and may require long and expensive transfers, a robust, affordable device with minimal storage may assist with risk stratification and triaging decisions. Furthermore, biomarker testing at pitch side may allow for the identification of sports-related concussion following head impact, allowing appropriate withdrawal of players (O’Brien et al. 2025). Even within the ED, an LFT device may offer an efficient method for sampling of GFAP at the initial triage to direct care pathways and timely resource allocation. Overall, a GFAP LFT, such as the LVOne, could serve as a valuable adjunct or alternative to current available analysis platforms, particularly in settings where simplicity, portability, and robustness are paramount. However, significant future work and evidence, including both validation studies and research to prove clinical effect, are required prior to clinical use.

A notable limitation of the LVOne GFAP LFT is the subjective nature of the scoring against the colorimetric score card, which inherently lacks the precision of fully quantitative assays and may pose challenges for clinical interpretation. The requirement for inter-observer agreement in this analysis highlights this limitation and reflects potential variability in score assignment. Digital reading approaches have previously been trialled for a variety of LFT tests, and would present a useful adjunct to this technology, improving accuracy and clinical governance. For example, digital readers have been shown to have a higher accuracy than human-read LFT HIV tests in rural South Africa (Turbé et al. 2021), and for diagnosis of SARS-CoV-2 during the COVID-19 pandemic (UK Health Security Agency 2022). A digital reader does not necessarily require a separate medical device, with smartphones commonly used to provide digital reading through custom made mobile phone applications (UK Health Security Agency 2022). A further limitation of the LVOne, outside of the controlled research environment, is the requirement to read the test at 15 min, with longer incubation times potentially invalidating the result. In time-pressured environments it may be difficult to strictly regulate time, and further investigation into a safe reading window may be required.

This study has additional limitations, the most notable being a retrospective analysis of a comparatively small sample size derived from a single disease state. The sample size was convenience based and limited by the availability of the LVOne LFTs, which leaves the study underpowered for true interrogation of the manufacturer reported lower limit of detection leading to large confidence intervals. Samples were selected based on the known quantified GFAP levels. Whilst this approach was intentionally designed to interrogate the LVOne GFAP LFA’s dynamic range, it may introduce selection bias and limit the generalisability of findings to an unselected clinical population. Ongoing prospective analysis of the LVOne is currently being conducted in a clinical stroke population. However, prior to clinical translation for different disease states, including TBI, further clinical evaluation is required.

The TBI patients studied were all admitted to neurocritical care, with the majority having a severe TBI. While this study was a technical examination of the LVOne GFAP LFA, it is important to note that the test may be of particular clinical value in the mild TBI population, and the lack of patients with mild TBI limits how far our results can be extrapolated clinically to this population. Therefore, targeted examination of the LVOne GFAP LFT within a mild TBI population is required to establish the utility and generalisability of this device. The lower limit of detection of the LVOne LFA (GFAP ≥ 0.2 ng/mL), as set by the manufacturer, is high in relation to the clinical thresholds of GFAP reported in the prior literature for TBI diagnosis or CT decision-making (Bazarian et al. 2018; Reyes et al. 2023; Papa et al. 2024). Although beyond the scope of this analysis, further assessment is required to determine the optimum cutoff of the LFA depending on the exact clinical use, with the lower limit of detection of the LVOne likely to require lowering for clinical diagnostic use in a TBI population. This may improve the accuracy of the test, particularly for mild TBI cases or regarding the diagnosis of sports-related concussion, where clinically important GFAP elevations are seen below the current detection threshold of the test (Reyes et al. 2023; Papa et al. 2024; O’Brien et al. 2025). The Quanterix Simoa® Human Neurology 4-Plex B assay is a research-use-only platform and is not intended for clinical diagnostic procedures. In this analysis, it was used as the reference comparator due to its analytical sensitivity, especially at low protein concentrations (Krausz et al. 2021), and widespread use in TBI research (Czeiter et al. 2020). Additionally, prior validation of the LVOne GFAP LFA has been conducted in a stroke population against a commercial PoC platform (Abbott iSTAT TBI Plasma), demonstrating a similar degree of correlation to that observed in our study (Gaude et al. 2025). However, future clinical validation of the LVOne in a TBI population should be undertaken in reference to clinically approved platforms.

There are multiple potential confounders of the acute GFAP level following TBI, including age, underlying comorbidities, and, critically, the timing of sample collection in relation to injury (Abdelhak et al. 2022). These factors can influence the interpretation of GFAP concentrations, and the thresholds considered clinically meaningful. However, the primary aim of this study was to compare the performance of the LVOne GFAP LFA with that of the gold-standard assay, focusing on analytical agreement rather than clinical interpretation. As such, biological and temporal determinants of GFAP expression, including sample timing, were not considered in this analysis. Future evaluations of the clinical utility of the LVOne will need to account for, and appropriately manage, these variables. Owing to the limited LVOne LFT availability, the samples were measured once without repeat testing, and therefore test–retest variability was not evaluated in this analysis. This represents a limitation and warrants further investigation in future studies. Finally, all LVOne LFT scoring was performed concurrently by observers, with a score only recorded following discussion, preventing the assessment of inter-observer reliability. Future evaluations of the LVOne should include independent observer scoring and formal assessment of inter-rater agreement.