Methods for enhancing specificity and sensitivity of group a
The present disclosure provides methods and kits for detecting Group A Streptococcus in biological samples. More particularly, the present disclosure provides methods for enhancing the specificity and sensitivity of Group A Streptococcus immunoassays by including N-propionyl-D-glucosamine, 2-N-butanoyl-D-glucosamide, Bis-(2-(D-2-deoxy-glucosaminyl))-PEG3-amide, m-PEG4-glucosamine, m-PEG6-glucosamine, or m-PEG10-glucosamine. The methods and kits disclosed herein are thus useful for reliable and early diagnosis of streptococcal infections in a subject.
1 . A device for detecting the presence of Group A streptococcus in a sample, comprising: a matrix having
(i) a sample receiving zone for receiving a sample containing or suspected of containing a Group A streptococcus -specific antigen,
(ii) a labeling zone containing a detector antibody for specifically labeling the antigen as it passes there through and
(iii) a capture zone having means for specifically binding the labeled antigen thereon, wherein the sample receiving zone, the labeling zone and the capture zone are arranged on the matrix in a liquid flow path, and
wherein at least one of the sample receiving zone, capture zone, and the labeling zone comprise N-propionyl-D-glucosamine or Bis-(2-(D-2-deoxy-glucosaminyl))-PEG3-amide,
wherein the detector antibody comprises a polyclonal antibody that specifically targets a Group A streptococcus -specific antigen, and
wherein the amount of N-propionyl-D-glucosamine, or Bis-(2-(D-2-deoxy-glucosaminyl)-PEG3-amide is an amount effective to reduce a false positive signal in the presence of mucin in the sample.
2 . The device of claim 1 , the detector antibody comprises a gold nanoparticle label, a chemiluminescent label, a radioactive label, a bioluminescent label, a fluorescent label, a chromogenic label, a spectroscopic label, a photochemical label, or an electrochemiluminescent label.
3 . The device of claim 1 , wherein the means for specifically binding the labeled antigen to the capture zone is a capture antibody or capture antigen binding fragment.
4 . The device of claim 3 , wherein the capture antibody is a polyclonal antibody.
5 . The device of claim 2 , wherein the fluorescent label is selected from the group consisting of europium, TagBFP, Azurite, EBFP2, mKalama1, Sirius, Sapphire, T-Sapphire, ECFP, Cerulean, SCFP3A, mTurquoise, monomeric Midoriishi-Cyan, TagCFP, mTFP1, EGFP, Emerald, Superfolder GFP, Monomeric Azami Green, TagGFP2, mUKG, mWasabi, EYFP, Citrine, Venus, SYFP2, Tag YFP, Monomeric Kusabira-Orange, mKOK, mKO2, mOrange, mOrange2, mRaspberry, mCherry, dsRed, mStrawberry, mTangerine, tdTomato, TagRFP, TagRFP-T, mApple, mRuby, mPlum, HcRed-Tandem, mKate2, mNeptune, NirFP, TagRFP657, IFP1.4, iRFP, mKeima Red, LSS-mKate1, LSS-mKate2, PA-GFP, PAmCherryl, PATagRFP, Kaede (green), Kaede (red), KikGR1 (green), KikGR1 (red), PS-CFP2, PS-CFP2, mEos2 (green), mEos2 (red), PSmOrange, and Dronpa;
the chemiluminescent label is selected from the group consisting of β-galactosidase, horseradish peroxidase (HRP), and alkaline phosphatase;
the bioluminescent label is selected from the group consisting of aequorin, firefly luciferase, Renilla luciferase, red luciferase, luxAB, or nanoluciferase;
wherein the radioactive label is selected from the group consisting of 3 H, 14 C, 32 P, 35 S, 36 Cl, 57 Co, 131 I and 186 Re.
6 . The device of claim 1 , wherein the sample is urine, saliva, sputum, mucous, a throat swab sample, whole blood, plasma, serum, amniotic fluid, semen, wound secretions, vaginal secretions, tears, or spinal fluid.
7 . The device of claim 1 , wherein the Group A streptococcus -specific antigen is selected from the group consisting of capsular polysaccharide (C-substance), cell wall peptidoglycan, lipoteichoic acid (LTA), M protein, fimbrial proteins, fibronectin-binding proteins, and cell-bound streptokinase.
8 . The device of claim 1 , wherein the sample is a human sample.
9 . The device of claim 1 , wherein the Group A Streptococcus species is Streptococcus pyogenes or Streptococcus dysgalactiae.
10 . The device of claim 1 , wherein the device comprises a lateral flow immunoassay device.
11 . The device of claim 10 , wherein the sample receiving zone comprises N-propionyl-D-glucosamine or Bis-(2-(D-2-deoxy-glucosaminyl))-PEG3-amide.
12 . The device of claim 10 , wherein the labeling zone comprises N-propionyl-D-glucosamine or Bis-(2-(D-2-deoxy-glucosaminyl))-PEG3-amide.
13 . The device of claim 1 , wherein the sample comprises epithelial cells originating from the tongue, cheek or teeth of a subject from whom the sample was obtained.
14 . The device of claim 1 , wherein the N-propionyl-D-glucosamine or Bis-(2-(D-2-deoxy-glucosaminyl))-PEG3-amide does not increase false negative results when the device is used.
15 . The device of claim 1 , wherein the sample is a human throat swab sample or human saliva sample, wherein the labeling zone comprises a polyclonal antibody that specifically targets a Group A streptococcus capsular polysaccharide (C-substance), wherein the polyclonal antibody comprises a gold nanoparticle label, and wherein the sample receiving zone or labeling zone comprises an amount of N-propionyl-D-glucosamine effective to reduce a false positive signal in the presence of mucin in the sample.