DOI: 10.5061/dryad.v15dv4239
title: Data from: Illuminating T cell-dendritic cell interactions in vivo by FlAsHing antigens

description: Delineating the complex network of interactions between antigen-specific T
 cells and antigen presenting cells (APCs) is crucial for effective
 precision therapies against cancer, chronic infections, and autoimmunity.
 However, the existing arsenal for examining antigen-specific T cell
 interactions is restricted to a select few antigen-T cell receptor pairs,
 with limited in situ utility. This lack of versatility is largely due to
 the disruptive effects of reagents on the immune synapse, which hinder
 real-time monitoring of antigen-specific interactions. To address this
 limitation, we have developed a novel and versatile immune monitoring
 strategy by adding a short cysteine-rich tag to antigenic peptides that
 emits fluorescence upon binding to thiol-reactive biarsenical hairpin
 compounds. Our findings demonstrate the specificity and durability of the
 novel antigen-targeting probes during dynamic immune monitoring in vitro
 and in vivo. This strategy opens new avenues for biological validation of
 T-cell receptors with newly identified epitopes by revealing the behavior
 of previously unrecognized antigen-receptor pairs, expanding our
 understanding of T cell responses.
description: The dataset was collected from flow cytometry and dynamic two
 photon imaging. The raw data are being deposited here.
description: # Illuminating T cell-dendritic cell interactions in vivo by FlAsHing
 antigens Give a brief
 summary of dataset contents, contextualized in experimental procedures and
 results. ## Description of the data and file structure The flow cytometry
 and imaging data for the Figures 1-6 are deposited here as fcs, ims and
 pzfx files. The files are: | Filename | File type | Figure  | Experimental
 Condition | | :------------------------------------------------- |
 :-------- | :------ | :---------------------- | | 6UBA70\_1.FCS | fcs | 1b
 | OVA | | 6U1A28\_1.FCS | fcs | 1b | C variant | | 6UMFLA\_2.FCS | fcs |
 1b | N variant | | 6U353A\_1.FCS | fcs | 1b | CACA | | 6UEB09\_1.FCS | fcs
 | 1b | NACA | | 6UB6C3\_1.FCS | fcs | 1b | DC-unpulsed | | FL0877\_1.FCS |
 fcs | 1c | No Antigen | | FL59C9\_1.FCS | fcs | 1c | Native peptide | |
 FL6FB4\_1.FCS | fcs | 1c | Variant peptide | | FL5A50\_1.FCS | fcs | 1c |
 No Antigen + TCEP | | FL528B\_1.FCS | fcs | 1c | Native peptide + TCEP | |
 FL5FAA\_1.FCS | fcs | 1c | Variant peptide + TCEP | |
 proliferation\_day\_3\_Tube\_001\_001.fcs | fcs | 2a-d | No antigen | |
 proliferation\_day\_3\_Tube\_015\_015.fcs | fcs | 2a-d | C-variant | |
 proliferation\_day\_3\_Tube\_014\_014.fcs | fcs | 2a-d | Native peptide |
 | proliferation\_day\_3\_Tube\_018\_018.fcs | fcs | 2a-d | N-variant | |
 proliferation\_day\_3\_Tube\_020\_020.fcs | fcs | 2a-d | CACA-variant | |
 proliferation\_day\_3\_Tube\_022\_022.fcs | fcs | 2a-d | NACA-variant | |
 No\_antigen-2.fcs | fcs | 2f | No Antigen | | OVA-2.fcs | fcs | 2f | OVA |
 | OVACACA-2.fcs | fcs | 2f | OVACACA | | b-r\_coloc\_FOR\_FLOWJO.fcs | fcs
 | 3g | OTII | | m-r\_coloc\_FOR\_FLOWJO.fcs | fcs | 3g | Polyclonal | |
 R\_Sec\_2\_1h\_20min\_.ims | ims | 5d | Refer to the figure | |
 MS1WHO\_1.IMS | ims | 4g | MHCII KO + WT | | MS2WHO\_1.IMS | ims | 4g |
 MHCI/II DKO + WT | | WTVSMH\_1.FCS | fcs | 3e | MHCII KO + WT | |
 Flash\_MS\_Fig\_4.pzfx | pzfx | 3f | analysis file | | WTVSMH\_2.FCS | fcs
 | 3e | MHCI/II DKO + WT | | OVA-Biotin\_021318\_Tube\_003\_003.fcs | fcs |
 4b | Refer to the figure | |
 OTII\_in\_vivo\_Day\_1\_030818\_Tube\_005\_015.fcs | fcs | 3b | Refer to
 the figure | | OTII\_in\_vivo\_Day\_2\_030918\_Tube\_005\_005.fcs | fcs |
 3b,c | Refer to the figure | | FlAsH-ReAsH\_122418\_Tube\_003\_009.fcs |
 fcs | 1d | Native peptide | | FlAsH-ReAsH\_122418\_Tube\_009\_015.fcs |
 fcs | 1d | Variant peptide | | FlAsH-ReAsH\_122418\_Tube\_006\_012.fcs |
 fcs | 1d | Native peptide + ReAsH | |
 FlAsH-ReAsH\_122418\_Tube\_012\_018.fcs | fcs | 1d | Variant peptide +
 ReAsH | | FlAsH-ReAsH\_122418\_Tube\_005\_011.fcs | fcs | 1e | native
 peptide | | FlAsH-ReAsH\_122418\_Tube\_011\_017.fcs | fcs | 1e |
 CACA-variant | | FlAsH-ReAsH\_122418\_Tube\_017\_023.fcs | fcs | 1e |
 CACA-2xTetracysteine | | FlAsH-ReAsH\_122418\_Tube\_023\_029.fcs | fcs |
 1e | CACA-BAL resistant | | PAPERF\_1.PZF | pzf | 2g | analysis file | |
 041819\_imaging-\_PAPER\_3e-h.pzfx | pzfx | 3 e,h | analysis file | |
 FL9A8D\_1.FCS | fcs | 4c | Refer to the figure | | MHC\_KO\_imaging.pzfx |
 pzfx | 4h | analysis file | | EAE873\_1.FCS | fcs | 4b | No antigen | |
 EAE1C0\_1.FCS | fcs | 4b | OVACACA | | EA86CC\_1.FCS | fcs | 4b | OVACACA
 + Eα(52-68) | | Flash\_MS\_Fig\_5.pzfx | pzfx | 4c | analysis file | |
 PAPER-Fig5\_e-f.pzfx | pzfx | 5e,f | analysis file | |
 Specimen\_001\_Tube\_002\_009.fcs | fcs | 6c-e | No peptide | |
 Specimen\_001\_Tube\_004\_011.fcs | fcs | 6c-e | OVA | |
 Specimen\_001\_Tube\_009\_016.fcs | fcs | 6c-e | OVACACA | |
 Specimen\_001\_Tube\_011\_018.fcs | fcs | 6c-e | Variant 1 | |
 Specimen\_001\_Tube\_013\_020.fcs | fcs | 6c-e | Variant 2 | |
 Specimen\_001\_Tube\_016\_023.fcs | fcs | 6c-e | Variant 3 | |
 FINAL\_Fig\_6.pzfx | pzfx | 6d,e | analysis file | ## Code/Software Flow
 Jo is the recommended software for the fcs files, Imaris is the
 recommended software for imaging files. Graphpad prism is the ideal
 software to view pzfx files.