Rapid Identification of 2009 H1N1 Influenza A Virus Using Fluorescent Antibody Methods

Abstract

Identification of the 2009 H1N1 influenza A virus requires emergency use authorized (EUA) molecular reverse transcriptase-polymerase chain reaction. Laboratories lacking molecular capabilities outsource testing, which is costly and may delay result reporting. A fluorescent antibody (FA; D(3) Ultra 2009 H1N1 influenza A virus ID Kit, Diagnostic Hybrids, Athens, OH) recently received Food and Drug Administration EUA status for 2009 H1N1 virus identification. The performance of this FA reagent was evaluated in this study. Influenza A-positive nasopharyngeal specimens (seasonal H1, H3, and 2009 H1N1) were prepared for culture and FA testing and were stained using influenza A antibodies and the 2009 H1N1 reagent. Other respiratory viruses were also evaluated. The FA reagent demonstrated 100% sensitivity and specificity. Bright, apple-green fluorescence was effortlessly identified in culture-positive cells, particularly around cell membrane perimeters. Laboratory-prepared slides were preferred over bedside-prepared specimens because background fluorescence obscured identification in the latter. The new FA reagent provides an accurate, rapid, and inexpensive assay for identifying the 2009 H1N1 virus in nonmolecular diagnostic laboratories.

Evaluation of Indirect Fluorescent Antibody Assays Compared to Rapid Influenza Diagnostic Tests for the Detection of Pandemic Influenza A (H1N1) pdm09

“…There is an IFA specifically for the diagnosis of A(H1N1)pdm09 that was approved by the Food and Drug Administration on an emergency use authorization basis, but it has only been evaluated in the lab [17], [18]. In hospitalized children, our IFA performed no better than RIDT.…”

“…Sensitivity of DFA for the detection of A(H1N1)pdm09 has been reported between 47 to 93% [1], [3], [6], [9], [11]–[16]. Information is not currently available on the accuracy of IFA for the detection of A(H1N1)pdm09 in a clinical setting, although a newly developed H1N1-specific IFA claims to have up to 100% sensitivity [17], [18]. More information on the accuracy of IFA tests compared to other diagnostic methods for the detection of A(H1N1)pdm09 is needed to determine their utility in the diagnosis and management of patients with febrile respiratory infections.…”

Evaluation of Alere i Influenza A&B for Rapid Detection of Influenza Viruses A and B

“…However, their sensitivities vary widely depending on the manufacturer and can be as low as 10%, with specificities ranging from 90 to 100% (14). DFAs are more sensitive than RIDTs and can be accomplished within 3 h but require skilled technologists for correct interpretation of results (15,16). Similarly, culture has increased sensitivity over both RIDTs and DFAs but requires skilled technologists and specialized laboratory settings and has a long turnaround time (2 to 14 days) (17).…”

Rapid Molecular Detection and Differentiation of Influenza Viruses A and B

“…Rapid influenza antigen detection tests (RIDTs) are widely used in clinical practice as point-of-care tests because they are simple to use and provide results within 15 to 30 min 11,12 ; however, their sensitivities vary widely (10-80%) depending on the manufacturer and the population being tested, and the influenza type and subtype 13,14,15 . Direct fluorescence assays (DFAs) provide superior sensitivities over RIDTs, but the processing time is greater (~3 hr) and must be completed by skilled technologists 16,17 . Viral culture has been the gold standard for influenza diagnostics and has improved sensitivity over both RIDTs and DFAs…”