Technical Note: Bimodal Negative Interference of Biotin in Roche IL-6 Immunoassay.

OBJECTIVE: Interleukin -6 (IL-6) is an important diagnostic test in COVID-19 patients to determine whether to initiate tocilizumab therapy or mechanical ventilation. We investigated potential interference of biotin in Roche IL-6 assay which utilizes biotinylated antibody. METHODS: We prepared three serum pools from left-over specimens which showed IL-6 values over 40 pg/mL. Then aliquots of each serum pool were further supplemented with various amounts of biotin expected in patients taking biotin supplement and then IL-6 values were measured again using Roche IL-6 assay on the Cobas e411 analyzer. RESULTS: We observed negative interference of biotin in IL-6 assay but interference was bimodal as maximum negative interference was observed with 100 ng/mL biotin but not with 1000 ng/mL. However, no interference was observed in the presence of 25 ng/mL biotin. CONCLUSIONS: Biotin showed negative interference with IL-6 assay.

Taking Advantage of Assay Harmonization, Biotin Interference in the LOCI Digoxin Assay Could Be Eliminated by Using the ADVIA Centaur Digoxin Assay.

Biotin at elevated concentration interferes with immunoassays that utilize biotin in assay design. We earlier reported interference of biotin in the luminescent oxygen channeling assay (LOCI) digoxin assay which utilizes biotinylated antibody against digoxin. However, the ADVIA Centaur digoxin assay, also manufactured by Siemens Diagnostics, does not utilize biotin in assay design. We hypothesized that if the LOCI and the ADVIA Centaur digoxin assay are harmonized, then interference of biotin in the LOCI digoxin assay could be eliminated by using the ADVIA Centaur digoxin assay. We analyzed 25 specimens from patients receiving digoxin using both assays to investigate harmonization between these two assays. Then aliquots of drug-free serum pool were supplemented with various biotin concentrations (range: 10 ng/mL to 2000 ng/mL) followed by measuring apparent digoxin levels using the ADVIA Centaur digoxin assay. In another set of experiments, aliquots of a serum digoxin pool were supplemented with biotin (10-2000 ng/mL) and digoxin concentrations were measured by the ADVIA Centaur digoxin assay. We observed an excellent correlation between digoxin values obtained by the LOCI digoxin assay (reference method) and the ADVIA Centaur digoxin assay (y= 1.0514 x+0.1083, r=0.99) indicating that both assays are harmonized. We did not observe any interference of biotin even at a highly elevated concentration of 2000 ng/mL with the ADVIA Centaur digoxin assay. We conclude that taking advantage of assay harmonization, interference of biotin in the LOCI digoxin assay can be eliminated by using the ADVIA Centaur digoxin assay.

The new LOCI digoxin assay on the Vista 1500 analyzer is virtually free from interferences of herbal supplements hawthorn and ashwagandha (Indian ginseng).

Herbal supplements hawthorn and ashwagandha (Indian ginseng) are indicated for cardiac illnesses and may be taken by patients receiving digoxin therapy. Because both hawthorn and ashwagandha are known to interfere with serum digoxin measurements using certain digoxin immunoassays, we investigated potential interference of these two herbal supplements with the new homogenous sequential chemiluminescent assay for digoxin based on the luminescent oxygen channeling technology (LOCI digoxin) for application on the Dimension and Vista platform. When aliquots of a drug-free serum pool were supplemented with various amounts of hawthorn (three different commercial preparations) or ashwagandha (two different commercial preparations) and apparent digoxin values were measured using LOCI digoxin assay on Dimension Vista 1500 analyzer we observed none-detected values except when aliquots were supplemented with very high amounts of the herbal extracts. When aliquots of a serum digoxin pool (prepared by pooling specimens from patients receiving digoxin) where further supplemented with various amounts of these supplements and digoxin concentrations were remeasured, statistically significant falsely higher digoxin values were observed only in specimens containing very high amounts of these supplements. Such interference may not be clinically significant. We conclude that new LOCI digoxin assay is virtually free from interferences of herbal supplements, hawthorn, and ashwagandha.

Impaired protein binding of Chinese medicine DanShen in uremic sera and sera with hyperbilirubinemia: rapid assessment of total and free DanShen concentrations using the fluorescence polarization immunoassay for digoxin.

DanShen is a Chinese medicine that is used to treat cardiovascular disorders. DanShen is moderately to strongly protein bound, mainly to albumin. Because impaired protein binding of albumin-bound drugs in uremia has been reported, we studied protein binding of DanShen by measuring the digoxin-like immunoreactive component of this Chinese medicine. We observed a significantly higher percentage of free fraction of DanShen in uremic sera in vitro. Impaired protein binding of DanShen was also observed in sera from patients with liver disease, who had elevated concentrations of bilirubin. Treating uremic sera with activated charcoal significantly improved the protein binding of DanShen, indicating that uremic compounds are responsible for the impaired protein binding of DanShen. On the other hand, when various amounts of bilirubin were added to aliquots of the normal pool supplemented with DanShen, we observed only a modest displacement of DanShen from the protein-binding sites by bilirubin, indicating that hypoalbuminemia may play a major role in impaired protein binding of DanShen in sera with elevated bilirubin concentrations. We conclude that protein binding of DanShen is lower in uremic sera and in sera with elevated bilirubin concentrations.

Activated charcoal is more effective than equilibrium dialysis in removing Chinese medicines Chan Su and Dan Shen from serum and activated charcoal also prevents further absorption of these agents from G.I. tract in mice: monitoring the effect in clinical laboratory by measuring digoxin activity in serum.

BACKGROUND: Chinese medicines are freely available without prescription and are widely used by the general population. Chan Su and Dan Shen are both indicated for the treatment of cardiac diseases. Severe toxicity from Chan Su has been reported. We studied the possibility of removing Chan Su and Dan Shen from human sera using activated charcoal and equilibrium dialysis, and also examined the potential benefit of preventing absorption of these agents from the G.I. tract in the mouse model. METHODS: For in vitro studies, drug-free serum pools were supplemented with Chan Su or Dan Shen and then either treated with activated charcoal (10 and 25 mg/ml), or passed through a column packed with activated charcoal. Serum pools supplemented with Chan Su or Dan Shen were also subjected to equilibrium dialysis against phosphate buffer (pH 7.4) using dialysis membrane with molecular cut-off of 25,000 Da. Removal of Chan Su or Dan Shen from the serum was monitored by measuring the apparent digoxin concentration using the fluorescence polarization immunoassay (FPIA) for digoxin (Abbott Laboratories). RESULTS: We observed the fast and effective removal of both Chan Su and Dan Shen from the serum by activated charcoal. We also observed significant removal of both Chan Su and Dan Shen when the serum pools containing these Chinese medicines were passed through columns packed with activated charcoal. Although equilibrium dialysis was also effective in removing these Chinese medicines from the serum, 24 h was required for complete removal of Dan Shen activity, and for Chan Su, complete removal was not achieved even after 24 h. In our in vivo model, we observed significantly less digoxin activity in the group of mice that received activated charcoal compared to the control group. CONCLUSIONS: Activated charcoal is effective in preventing absorption of these Chinese medicines from the G.I. tract and can also remove these agents from the serum.