The use of nanobodies in a sensitive ELISA test for SARS-CoV-2 Spike 1 protein.
The use of nanobodies in a sensitive ELISA test for SARS-CoV-2 Spike 1 protein.
复制标题
纳米抗体在SARS-CoV-2 Spike 1蛋白的灵敏ELISA测试中的用途。
DOI：
10.1098/rsos.211016
复制
发表时间：
2021-09
影响因子：
3.5
通讯作者：
Naismith JH
中科院分区：
文献类型：
--
作者：
Girt GC;Lakshminarayanan A;Huo J;Dormon J;Norman C;Afrough B;Harding A;James W;Owens RJ;Naismith JH
检测体液中的严重急性呼吸综合征冠状病毒2（SARS-CoV-2）抗原在生物技术中具有重要用途，并且是许多护理点SARS-CoV-2诊断的组成部分。夹心酶联免疫吸附试验（ELISA）是一种灵敏、成熟的测定溶液中抗原的方法。它们使用一种配体捕获目标分析物，另一种配体检测目标分析物。检测通常使用在底物与HRP缀合的第二配体反应后获得的比色读数来实现。纳米抗体，骆驼抗体的VHH结构域，已经扩大了抗原检测中使用的分子库。纳米抗体对靶抗原的高亲和力、其紧凑的结构、其高稳定性和易于生产已经推动了对其用作诊断试剂的研究。在对SARS-CoV-2刺突蛋白受体结合结构域上的表位的结构理解的指导下，我们在夹心ELISA中研究了工程化纳米抗体的各种组合以检测SARS-CoV-2的刺突蛋白。我们已经确定了纳米抗体的最佳组合。这些被选择性地官能化以进一步改善抗原捕获，使得能够测量溶液中亚皮摩尔量的SARS-CoV-2刺突蛋白。在这种组合下，通过加热和洗涤剂灭活的样品中的常规检测限对应于小于7个传染性SARS-CoV-2的病灶形成单位。
Detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antigens in the fluid has important uses in biotechnology, and is integral to many point-of-care SARS-CoV-2 diagnostics. Sandwich enzyme-linked immunosorbent assays (ELISAs) are a sensitive, well-established method of measuring antigens in solutions. They use one ligand to capture and the other ligand to detect the target analyte. Detection is commonly achieved using colorimetric readout obtained upon the reaction of a substrate with HRP-conjugated secondary ligand. Nanobodies, the VHH domain of camelid antibodies, have expanded the repertoire of molecules used in antigen detection. Nanobodies' high affinity for target antigens, their compact structure, their high stability and ease of production has driven research into their use as diagnostic reagents. Guided by a structural understanding of epitopes on the receptor-binding domain of the SARS-CoV-2 Spike protein, we investigated various combinations of engineered nanobodies in a sandwich ELISA to detect the Spike protein of SARS-CoV-2. We have identified an optimal combination of nanobodies. These were selectively functionalized to further improve antigen capture, enabling the measurement of sub-picomolar amounts of SARS-CoV-2 Spike protein in solution. With this combination, the routine detection limit in samples inactivated by heat and detergent corresponded to less than seven focus-forming units of infectious SARS-CoV-2.
登录
查看更多内容
影响因子：
4.6
作者：
Afrough B;Eakins J;Durley-White S;Dowall S;Findlay-Wilson S;Graham V;Lewandowski K;Carter DP;Hewson R
通讯作者：
Hewson R
DOI：
10.1016/s0140-6736(20)32661-1
发表时间：
2021-01-09
期刊：
Lancet (London, England)
影响因子：
--
作者：
Voysey M;Clemens SAC;Madhi SA;Weckx LY;Folegatti PM;Aley PK;Angus B;Baillie VL;Barnabas SL;Bhorat QE;Bibi S;Briner C;Cicconi P;Collins AM;Colin-Jones R;Cutland CL;Darton TC;Dheda K;Duncan CJA;Emary KRW;Ewer KJ;Fairlie L;Faust SN;Feng S;Ferreira DM;Finn A;Goodman AL;Green CM;Green CA;Heath PT;Hill C;Hill H;Hirsch I;Hodgson SHC;Izu A;Jackson S;Jenkin D;Joe CCD;Kerridge S;Koen A;Kwatra G;Lazarus R;Lawrie AM;Lelliott A;Libri V;Lillie PJ;Mallory R;Mendes AVA;Milan EP;Minassian AM;McGregor A;Morrison H;Mujadidi YF;Nana A;O'Reilly PJ;Padayachee SD;Pittella A;Plested E;Pollock KM;Ramasamy MN;Rhead S;Schwarzbold AV;Singh N;Smith A;Song R;Snape MD;Sprinz E;Sutherland RK;Tarrant R;Thomson EC;Török ME;Toshner M;Turner DPJ;Vekemans J;Villafana TL;Watson MEE;Williams CJ;Douglas AD;Hill AVS;Lambe T;Gilbert SC;Pollard AJ;Oxford COVID Vaccine Trial Group
通讯作者：
Oxford COVID Vaccine Trial Group
影响因子：
64.8
作者：
Barnes CO;Jette CA;Abernathy ME;Dam KA;Esswein SR;Gristick HB;Malyutin AG;Sharaf NG;Huey-Tubman KE;Lee YE;Robbiani DF;Nussenzweig MC;West AP Jr;Bjorkman PJ
通讯作者：
Bjorkman PJ
DOI：
10.1073/pnas.2014739117
发表时间：
2020-09-15
影响因子：
11.1
作者：
Ganguli A;Mostafa A;Berger J;Aydin MY;Sun F;Ramirez SAS;Valera E;Cunningham BT;King WP;Bashir R
通讯作者：
Bashir R
影响因子：
6.4
作者：
Koczula KM;Gallotta A
通讯作者：
Gallotta A