![]() ![]() The arrows indicate binding by the human monoclonal TPO autoantibody Fab WR1.7 (as a bacterial streak) that preferentially recognizes N-TPO. Controls include normal (Nl) human serum and normal mouse serum (e,f). In contrast, 125I-N-TPO is recognized by both serum antibodies and mouse MoAb 47 (f). 125I-DN-TPO used in these studies is recognized very well by mouse MoAb 47, but only weakly or not at all by serum autoantibodies (patients GA and FA) (e). Unexpectedly, DN-TPO binding clones interact better with 125I-N-TPO (c) than with 125I-DN-TPO (d). As anticipated, DN-TPO-selected clones bind 125I-DN-TPO (a) and N-TPO-selected clones bind 125I-N-TPO (b). Individual clones from the third round of panning against DN-TPO or N-TPO were tested in filter lifts for binding of radiolabelled antigen. Interaction of thyroid-derived clones with native (N) versus denatured (DN) thyroid peroxidase (TPO) after three rounds of panning a pComb3H phage display library constructed from thyroid tissue of a patient with autoimmune thyroid disease. Our findings demonstrate the bias of the human B cell repertoire towards recognition of an immunodominant region on the conformationally intact form of a major thyroid autoantigen. In conclusion, human monoclonal autoantibodies that preferentially recognize denatured TPO could not be isolated from an immunoglobulin gene library despite selection with denatured protein. Although DN4 binds specifically to TPO, its affinity is low, unlike the high affinities of other human TPO autoantibodies. One clone (DN4), isolated using DN-TPO, did not interact with the TPO immunodominant region and its H chain derives from a different VH gene. Moreover, regardless of selection with N-TPO or DN-TPO, their heavy and light chains were encoded by similar VDJ and Vkappa combinations. Of 13 clones selected using DN-TPO or N-TPO, 12 clones recognized the TPO immunodominant region. Surprisingly, the library enriched after panning on DN-TPO, as well as a panel of individual clones, preferentially bound native (N)-TPO. Thyroid-infiltrating B cells from a typical TPO autoantibody-positive patient were the source of mRNA for library construction. We therefore searched an immunoglobulin gene phage display library for human autoantibodies that bind TPO denatured by reduction and alkylation (DN-TPO). However, none of the TPO-specific, human monoclonal autoantibodies isolated to date preferentially recognize denatured autoantigen. Contrary data demonstrate TPO autoantibody reactivity with DN-TPO or polypeptide fragments. There is evidence that autoantibodies to thyroid peroxidase (TPO) interact with overlapping conformational epitopes in an immunodominant region and binding to denatured (DN) protein is decreased. NFTs are just like any other ERC20 token you may have sent in the past and can be viewed on Etherscan or accessed via other Ethereum supported wallets like Trust Wallet and Math Wallet.The nature of the autoantibody repertoire to the dominant autoantigen in human autoimmune thyroid disease is controversial. Once you’ve bought a CLONE X it is automatically stored in your Metamask wallet and can then be relisted or sent to another Ethereum address at any time. To buy a CLONE X, click on the one you wish to buy and click “Buy now”, OpenSea will then walk you through a set of gas fees that must be paid before the CLONE X is yours. OpenSea will automatically filter to the “lowest floor price” which includes current offers and asking prices for CLONE Xs. Nov 29, 11pm CET: Public Sale event: minting of the CloneX Mintvials 8'367 items Dutch Auction. Nov 2729: Pre-Sale event: minting of the CloneX Mintvials 11'133 items. Once you’re signed in, head over to the CLONE X page on OpenSea to browse the CLONE Xs that are for sale. Weeks before Nov 27: Mintdiscs airdrop for those eligible (c.f. Follow the prompts to sign into OpenSea with your Metamask. ![]() First, connect your Metamask wallet to OpenSea by clicking on the wallet icon in the top right of the page. Because OpenSea allows you to buy CLONE X on-chain, it’s important to understand functions and costs involved with buying CLONE Xs. ![]()
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