What methods detect spike protein persistence and how reliable are they?
Executive summary
Multiple laboratory approaches detect SARS‑CoV‑2 spike protein or spike‑specific immune responses: direct protein assays (ELISA, highly sensitive biosensors like SPR, aptasensors, electrochemical sensors) and indirect surrogate neutralization or antibody serology testing [1] [2] [3] [4] [5]. Reported analytical sensitivity ranges from pg/mL and below for advanced SPR or graphene‑quantum‑dot chips (0.01 pg/mL claimed) to standard ELISAs and surrogate neutralization assays that measure antibody binding rather than intact circulating spike [3] [5] [2].
1. What “detecting spike protein persistence” means in the lab — two different signals
Studies use two broad strategies: direct detection of spike antigen in blood or cells, and indirect detection via antibodies or functional neutralization assays. Direct antigen detection methods include ELISA on plasma and specialized biosensors to capture spike or S1/RBD fragments [1] [2]. Indirect approaches measure anti‑spike antibody levels and map their decay or functional capacity with surrogate neutralization tests; these track immune memory rather than ongoing circulating antigen [5] [6].
2. Direct protein detection: methods and claimed sensitivities
Conventional ELISA can detect spike in plasma and was used in clinical studies that reported spike in some PASC/long‑COVID cohorts [1]. Research biosensors push sensitivity far lower: nitrogen‑doped graphene quantum‑dot SPR chips reported a limit of detection of 0.01 pg/mL for RBD in plasma [3]. Other platforms include interdigitated microelectrodes with dielectrophoresis and aptamer‑based reusable aptasensors that convert spike binding into measurable electrical or optical signals [2] [4]. Analytical papers also describe electrochemical nanorod/AuNP platforms and nanosheet probes for rapid RBD detection in saliva [7] [8]. These techniques trade clinical validation for increased sensitivity and speed [3] [4].
3. Indirect detection: serology and surrogate neutralization — what they actually measure
Serology assays quantify anti‑spike antibodies; surrogate neutralization assays estimate functional blocking of spike‑ACE2 interaction rather than measuring spike antigen itself [5] [6]. Longitudinal cohort data showed >95% of health‑care workers had detectable S antibodies up to ~200 days after infection, but only a subset retain titers sufficient for measurable surrogate neutralization [5] [9]. Thus persistent seropositivity does not equal persistent circulating spike protein [5].
4. Reliability: sensitivity, specificity, and biological interpretation differ by method
High‑sensitivity biosensors may detect minuscule spike fragments but lack large‑scale clinical validation and can be prone to cross‑reactivity or matrix effects unless rigorously benchmarked [3] [2]. ELISA is widely used in clinical studies and easier to standardize, but its detection limit is higher and it may miss very low‑level antigenemia [1]. Serology and surrogate neutralization are reliable for prior exposure and immune dynamics, not for proving antigen persistence; they model antibody kinetics and functional capacity, not ongoing antigenemia [5] [6].
5. Conflicting findings and interpretive pitfalls in the literature
Some clinical reports find spike protein or S1 subunit in plasma or monocytes months after infection or vaccination using sensitive assays [1] [10]. Other work cautions that antibody persistence is common while detectable functional neutralization wanes for many people [5]. Available sources do not mention standardized thresholds that distinguish residual fragmented antigen from biologically active, replication‑competent viral protein; many studies acknowledge possible detection of fragments rather than intact, pathogenic spike [1] [5].
6. Practical takeaways for clinicians and researchers
To claim “spike persistence” requires assay choice transparency: report method (ELISA vs ddPCR‑coupled antigen assays vs SPR/aptasensor), limit of detection, sample handling (e.g., RNase treatment for RNA, plasma vs cell fractions), and validation against known negatives/positives [1] [3] [4]. Use serology and surrogate neutralization to assess immune memory and use high‑sensitivity antigen assays cautiously when inferring pathophysiological relevance [5] [6] [1].
Limitations: sources focus on method development and cohort studies; large‑scale clinical validation and standardized clinical cutoffs for “persistence” are not established in the provided material (available sources do not mention standardized clinical thresholds).