Rapid detection of proteins by enzyme‐linked immunofiltration assay after transfer onto nitrocellulose membranes

Abstract

Enzyme-linked immunofiltration assay (ELIFA) for labeling transferred proteins is an interesting and powerful technique for the rapid specific detection (15 min) of proteins immobilized on nitrocellulose or nylon membranes (0.20 and 0.45 micron). ELIFA does not require fastidious handling of the membranes. Saturation, specific labeling and washing procedures are achieved by filtration, controlled by a monitoring unit which regulates the flow rate and ensures excellent specificity, repetition and reproducibility. The recycling by closed circuit or by repetitive inversion of the flow direction offers the advantage of reducing the volumes of expensive reagents while simultaneously increasing the sensitivity of the technique. The detection limit is at least as low as 1-5 ng using directly or indirectly enzymatically labelled probes. ELIFA may be extended to the identification of glycoproteins using specific ligands such as lectins or to the immunocapture of an antigen using specific antibodies immobilized on an activated membrane. ELIFA complements fast separation, by e.g., isoelectric focusing, polyacrylamide gel electrophoresis, or sodium dodecyl sulfate-polyacrylamide gel electrophoresis and accelerated electrotransfer to membranes with rapid detection reducing the total time for separation transfer and detection to less than 2 h.

Polyelectrolyte multilayers-modified membrane filter for rapid immunoassay: protein condensation by centrifugal permeation

“…; Figure 1b); hence, the incubation time for the antigen-antibody reaction corresponded to just the permeation time, which was 1/20 shorten than that of conventional ELISA systems (37 1C 1 h) and (3) this rapid and precise centrifugal permeation system can be applied to various molecular diagnostic biosensors because of its simple, reliable and disposable process, which improves other approaches for permeation detection in existence. [13][14][15][16][17] Moreover, we considered that this permeation detection system can achieve the separation and detection of antigen molecules from whole blood simultaneously, just when the molecular cutoff of the filter is appropriate to the molecular weight of the antigen.…”

Recent Developments in Tuberculous Meningitis Pathogenesis and Diagnostics

“…In ventricular CSF of children with TBM, transcriptome analysis has revealed significant enrichment of transcripts associated with neuro-excititoxicity predominantly driven by glutamate release and NMDA binding and receptor uptake 33 . Upregulation of genes associated with nitric oxide, cytochrome c, brain injury proteins like myelin basic protein, and proteins including tau, amyloidbeta and apo-lipoprotein were also seen 33 ; many of which have also been described in in neurodegenerative conditions such as Alzhiemer's disease and traumatic brain injury 34 .…”

Accelerated Detection of DNA on Membranes by Automated Enzyme-Linked Immunofiltration Assay

“…This used this technique (which has previously been employed to detect antibodies and proteins immobilized probe (Burg's oligo 3 (17)), is labeled with digoxigenin by tailing with terminal transferase (11). Stringency is on membranes (15,16)) to detect Toxoplasma gondii DNA in 108 samples of amniotic fluid sent to us for carried out in four steps (i) 2 1 10 min in buffer 2 (21 SSC, 0.1% SDS) at room temperature, and (ii) 2 1 15 prenatal diagnosis of toxoplasmosis.…”

“…They were washed in buffer consisting of TrisThis technique, first described by Pinon, was initially HCl 10 mM, EDTA 1 mM to obtain a cell pellet free of developed for the rapid detection of proteins after hemoglobin. The DNA was extracted from the pellets transfer to a membrane (15,16,18). We adapted it to by alkaline lysis (Tween 20 0.5%, Nonidet P-40 0.5%, the detection of nucleic acids immobilized on a memNaOH 10 mM) for 10 min at 100ЊC.…”