Ultra-Sensitive and Semi-Quantitative Vertical Flow Assay for the Rapid Detection of Interleukin-6 in Inflammatory Diseases

Abstract

The inflammation biomarker Interleukin 6 (IL-6) exhibits a concentration of less than 7 pg/mL in healthy serum but increases 10–100-fold when inflammation occurs. Increased serum IL-6 has been reported in chronic diseases such as rheumatoid arthritis (RA), as well as in life-threatening acute illnesses such as sepsis and cytokine release syndrome (CRS). This work seeks to meet the demand for rapid detection of serum IL-6 both for rapid monitoring of chronic diseases and for triaging patients with acute illnesses. Following the optimization of several types of gold nanoparticles, membrane pore sizes, and buffer systems, an ultra-sensitive vertical flow assay (VFA) was engineered, allowing the detection of recombinant IL-6 in spiked buffer with a limit of detection (LoD) of 10 pg/mL and a reportable range of 10–10,000 pg/mL with a 15-min assay time. The detection of IL-6 in spiked pooled healthy serum exhibited an LoD of 3.2 pg/mL and a reportable range of 10–10,000 pg/mL. The VFA’s stability was demonstrated over 1-day, two-week, four-week, and six-week storage durations at room temperature. The inter-operator CV and intra-operator CV were determined to be 14.3% and 15.2%, respectively. Three reference zones, high, low, and blank, were introduced into the cartridge to facilitate on-site semi-quantitative measurements across a 6-point semi-quantitative range. Finally, the performance of the IL-6 VFA was validated using 20 RA and 20 healthy control (HC) clinical serum samples, using ELISA as the gold standard platform. The ultra-sensitive, rapid IL-6 VFA could potentially be used to triage patients for intensive care, treatment adjustments, or for monitoring disease activity in inflammatory conditions.

Duplex Vertical-Flow Rapid Tests for Point-of-Care Detection of Anti-dsDNA and Anti-Nuclear Autoantibodies

“…The VFA addresses these issues by utilizing downward flow. Gravity assists in the capillary action of fluids, which helps yield faster results when compared to those obtained under lateral flow [40,41].…”

Evaluating drug interaction potential from cytokine release syndrome using a physiologically based pharmacokinetic model: A case study of teclistamab

“…[4][5][6][7][8] Among cytokines released during CRS and other inflammatory states, IL-6 is the best known potent suppressor of CYP enzyme activity. 2,9 For example, in vitro studies have demonstrated that IL-6 incubated at physiological concentrations led to decreased expression or activity of CYP1A2, CYP2C9, CYP2C19, CYP3A4, and CYP3A5 in human hepatocytes. 3,10 Importantly, this change in CYP activity has been confirmed clinically in patients with RA, who present IL-6 systemic concentrations ~50 pg/mL, and with higher exposure in CYP substrates compared with healthy patients.…”

“…Immunologically active proteins including interferons (IFNs), interleukins (ILs), and other cytokines are produced in response to infection or inflammation, and their systemic levels are elevated in chronic and acute inflammatory conditions, such as rheumatoid arthritis (RA) or cytokine release syndrome (CRS) 1,2 . Cytokines have been shown to alter the activity of cytochrome P450 (CYP) enzymes by affecting their stability or transcription factor activity for CYP enzyme expression, including PXR and CAR 3,4 .…”

“…Among cytokines released during CRS and other inflammatory states, IL‐6 is the best known potent suppressor of CYP enzyme activity 2,9 . For example, in vitro studies have demonstrated that IL‐6 incubated at physiological concentrations led to decreased expression or activity of CYP1A2, CYP2C9, CYP2C19, CYP3A4, and CYP3A5 in human hepatocytes 3,10 .…”

IL-6 Mutation Attenuates Liver Injury Caused by Aeromonas hydrophila Infection by Reducing Oxidative Stress in Zebrafish

“…Under normal physiological conditions, the levels of IL-6 in tissues and plasma are typically low. However, IL-6 expression increases rapidly and substantially when stimulated by pathogenic microorganisms such as bacteria, fungi, and viruses [6]. Numerous studies have shown that overproduction of IL-6 triggers a cytokine storm that leads to excessive inflammation in the body [7].…”