Here is a list of protocols, SOPs and associated resources generated in our lab.
TDP-43 RNA aptamer immunohistochemical staining in FFPE human tissue
Context: Our TDP-43 RNA aptamer detects pathological TDP-43 with greater sensitivity and specificity than currently available TDP-43 antibodies revealing novel pathological features of ALS. Our findings are published here
Spence H*, Waldron FM*, Saleeb RS, Brown AL, Rifai OM, Gilodi M, Read F, Roberts K, Milne G, Wilkinson D, O’Shaughnessy J, Pastore A, Fratta P, Shneider N, Tartaglia GG, Zacco E, Horrocks MH, Gregory JM. RNA aptamer reveals nuclear TDP-43 pathology is an early aggregation event that coincides with STMN-2 cryptic splicing and precedes clinical manifestation in ALS. Acta Neuropathologica 2024 Mar 5;147(1):50 *equal contributions
Application: Our immunohistochemistry protocol allows for pathological TDP-43 staining in FFPE human tissue with higher sensitivity and specificity than with available antibodies.
SOP: Below we provide an SOP, with associated tick-sheet, for staining FFPE human tissue with TDP-43 RNA aptamer:
Waldron FM*, Spence H*, Gregory JM‡. TDP-43 RNA aptamer staining to detect pathological TDP-43 in FFPE human tissue, as described in Spence and Waldron et al., 2024 (Acta Neuropathologica): A SOP and tick-sheet. v2. protocols.io 2024; DOI: dx.doi.org/10.17504/protocols.io.eq2lyjo4mlx9/v2. *equal contributions, ‡corresponding author.
An adaptable protocol for antibody & TDP-43 RNA aptamer dual immunohistochemical staining in FFPE human tissue
Context: This adaptable protocol (with SOP and tick-sheet) is designed to provide the user with a resource to dual stain their antibody of interest and TDP-43 RNA aptamer in FFPE human tissue. As with all dual staining, we recommend that users optimise single stains first, before proceeding to dual staining. This protocol is available from protocols.io but has not, as yet, been included/employed in a peer-reviewed journal article.
Application: This protocol can be implemented for dual staining of any rabbit or mouse primary detection antibody (using the Novolink Polymer Detection System), with our TDP-43 RNA aptamer (Spence and Waldron, 2024). The resulting dual stain will show brown chromogen (detecting the target of the primary antibody) with red chromogen (detecting TDP-43 pathology with our TDP-43 RNA aptamer).
SOP: Below we provide an SOP, with associated tick-sheet, for dual staining with antibody and TDP-43 RNA aptamer.
Waldron FM‡, Gregory JM‡. An adaptable protocol for antibody & TDP-43 RNA aptamer dual immunohistochemical staining for FFPE-preserved human tissue: with SOP and tick-sheet. protocols.io 2024. DOI: dx.doi.org/10.17504/protocols.io.yxmvm97k6l3p/v1. ‡co-corresponding authors.
NEK1 antibody & TDP-43 RNA aptamer dual immunohistochemical staining in FFPE human tissue
Context: We present a protocol (with SOP and tick-sheet) to dual stain NEK1 ((NIMA-related kinase 1) antibody and TDP-43 RNA aptamer in FFPE human tissue. NEK1 has been associated with an increased risk of ALS.
Provided in the protocol are optimised conditions and reagent concentrations for NEK1 antibody and TDP-43 RNA aptamer dual staining. The protocol is available at protocols.io and was employed in this publication:
Rifai OM, Waldron FM, Sleibi D, O’Shaughnessy J, Leighton DJ*, Gregory JM*‡. Clinicopathological analysis of NEK1 variants in amyotrophic lateral sclerosis. Brain Pathology. 2024. e13287. DOI: 10.1111/bpa.13287, *equal contributions, ‡corresponding author.
Application: This protocol can be implemented for dual staining of Rabbit Anti-NEK1 antibody (AB186519)(using the Novolink Polymer Detection System), with our TDP-43 RNA aptamer (Spence and Waldron, 2024). The resulting dual stain will show brown chromogen (detecting the target of the primary antibody) with red chromogen (detecting TDP-43 pathology with our TDP-43 RNA aptamer).
SOP: Below we provide an SOP, with associated tick-sheet, for dual staining with NEK1 antibody and TDP-43 RNA aptamer.
Waldron FM, Rifai OM, Gregory JM‡. Antibody and TDP-43 RNA aptamer dual staining to detect patterns of co-pathology in FFPE-preserved human tissue, as described in Rifai et al., 2024 (Brain Pathology): An SOP and tick-sheet. protocols.io 2024. DOI: dx.doi.org/10.17504/protocols.io.14egn6wnml5d/v1. ‡corresponding author.
