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Antibodies that Work In Tribolium Castaneum

Antibodies that Work In Tribolium Castaneum. Kevin Burns 5/20/2011. Notes on this Document. For antibodies I’ve used personally, the titles contain the following information Original antigen used to produce antibody A rough guess at the stage/stages of Tc shown

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Antibodies that Work In Tribolium Castaneum

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  1. Antibodies that Work In Tribolium Castaneum Kevin Burns 5/20/2011

  2. Notes on this Document • For antibodies I’ve used personally, the titles contain the following information • Original antigen used to produce antibody • A rough guess at the stage/stages of Tc shown • Antibody procurement info, e.g. DSHB, Supplier, or Lab • Host species and working dilution • I have often thrown in co-stains to further illustrate morphology and to make things look cool. However, mostly this is just confusing. • KT817 is an enhancer trap line from Sue Brown that marks oenocytes. It is also cool for marking A1-A8 • The second half of this document in-exhaustively covers antibodies that have been used in published works, normally on beetle, but occasionally on other arthropods, please update me if you find any new ones (Kevin.Burns@cchmc.org) • There is a PDF and a Powerpoint version of this document (The hyperlinks work in the PPT), if your email server does not support attachments of such & such size, I will post this document on http://burnska08.wordpress.com/kevin-burns-home-page/

  3. Index • AbdA (+UBX) • AntP • Futsch (22c10) • HNF4 • Notch Intracellular Domain • Orthodenticle (GP / Rat) • Phospho-Histone3 • Phospho-MAPK • Spalt • Trachea (a2A12) • UBX (see AbdA) Index of Other’s Results

  4. Dm Abd-A/UBXPre-Hatching, ~64h at 30CDSHB: Mouse Monoclonal, 1:5 UBX/AbdA DAPI KT817-YFP UBX/AbdA Index for KB’s Antibodies

  5. Dm AntPGermband retraction 30h @30CDSHB: Mouse Monoclonal, 1:25+ KT817-YFP DAPI AntP AntP NICD Please advise naïve young investigators that AntP is not just expressed in thoracic segments but also in the CNS Index for KB’s Antibodies

  6. Dm En/InvDorsal closure/Leg articulation (36h)DSHB: 4D9Mouse Monoclonal, 1:25 En/Inv Spalt En/Inv Index for KB’s Antibodies

  7. Tc EngrailedCocalico/Gebelein, coming in JulyRat/Guinea Pig Kevin Burns is making this in Brian Gebelein’s lab in order to do some co-stains with the multitude of mouse antibodies Index for KB’s Antibodies

  8. Dm FutschDorsal closure/48h? (Ventral View)DSHB: 22c10Mouse Monoclonal, 1:50 22c10 Index for KB’s Antibodies

  9. HNF448h @ 30CCocalico/Gebelein: Rat Serum, 1:1000 KT817-YFP HNF4 p HNF4 Mesodermal staining (arrowheads), continues underneath more superficial oenocyte staining (e.g. red circle) p = pleuropod in A1 Index for KB’s Antibodies

  10. Notch Intracellular DomainGermband extensionDSHB: C17.9C6Mouse Monoclonal 1:5 NICD NICD DAPI Index for KB’s Antibodies

  11. Dm OTD full lengthMix sampleCocalico/Cook: Rat Serum, 1:500 OTD Index for KB’s Antibodies I am conducting another stain to see if this detects the OTD that replaces bicoid in early patterning…

  12. Dm OTD full lengthMix sampleCocalico/Cook: Guinea Pig Serum, 1:500 OTD Index for KB’s Antibodies

  13. phospho-Histone 3Full extensionMillipore: 05-806Mouse, 1:1000 pH3 pH3 DAPI Index for KB’s Antibodies

  14. Di-Phosphorylated MAPKpost-gastrulation and full germbandSigma: M-8159Mouse Monoclonal, 1:50 pERK pERK Index for KB’s Antibodies

  15. Dm Spalt Major24h, 36h @ 30CCocalico/Gebelein: Rabbit Serum, 1:1500 Spalt Spalt pERK Spalt Spalt KT-817-YFP Index for KB’s Antibodies

  16. Dm unknown (tracheal System)Pre-HatchingDSHB: a2A12 Mouse IGM Monoclonal, 1:25 This stain was achieved with IgG secondary antibodies, my guess is secondaries designed against IgM would fare much better Index for KB’s Antibodies NICD a2A12 DAPI

  17. Table of Kevin’s Antibodies Index for KB’s Antibodies

  18. Index of Other People’s • Caudal • Caspase3 • Cx (scr) • DC0(mushroom body etc) • DLL • engrailed 4F11 from DSHB • Even-skipped • Nubbin (Drosophila + Crustacean + Chelicerates) • Prd, Gooseberry, Gooseberry-Neuro (Pairberry) • pSMAD • pY • Synapsin 1 Index for KB’s Antibodies

  19. CaudalShulz&Tautz 1998 Dev Genes & Evolution Generation of an antibody Part of the cadA ORF (open reading frame, corresponding to amino acids 79–249) was amplified by PCR and cloned in frame into the EcoRV site of the pRSET-C (Invitrogen) vector. The Index of Other’s Results

  20. Caspase 3Aranda&Tautz2009 Dev Genes Evol For apoptosis detection using the anti cleaved caspase3 antibody (Cell Signalling Technology, Inc.) Index of Other’s Results

  21. Cx (Scr)Shippy & Denell 2006 Genetics A cross-reacting polyclonal antibody to Drosophila Scr, α-DmScr (a gift from Thomas Kaufman), was used to detect Cx Index of Other’s Results

  22. DC0Zhao&Farris 2008 Anti-DC0 primary antibody, a polyclonal antibody against the catalytic subunit of Drosophila melanogaster protein kinase A, was a gift provided by Dr. Daniel Kalderon. This antibody has been shown to have a high affinity for mushroom body intrinsic neurons in a range of insect species (Skoulakis et al., 1993; Farris, 2005a,b). Index of Other’s Results

  23. DLLe.g. Panganiban&Carroll 1997 PNAs • Butterfly Staining… Production and characterization of the Dll homeodomain antibody was described in Panganiban et al. (4) Index of Other’s Results

  24. Engrailed 4F11 (alternate DSHB Clone)e.g. Pueyo&Couso2008 PNAS This is cochroach, but I swear I’ve seen beetle people use 4f11 instead of 4D9 Index of Other’s Results

  25. Even SkippedBrown & Denell 1997 Choe & Brown 2006 Grasshopper antigen, see also Patel 1994 Index of Other’s Results

  26. NubbinLi & Popadic 2004 Evolution& Development A monoclonal antibody that recognizes a conserved epitope in the Nub/pdm protein has been generated and used successfully in a number of crustacean and chelicerate species (Averof and Cohen 1997; Abzhanov and Kaufman 2000; Damen et al. 2002). Unfortunately, this antibody does not recognize insect nub orthologs except for that of Drosophila. Index of Other’s Results

  27. Pairberry (Prd, Gsb, Gsbn)Davis & Patel 2001 Index of Other’s Results

  28. Phospho-SMADNunes da fonseca& Roth 2008 Dev Cell Index of Other’s Results

  29. Phospho-TyrosineNunes da fonseca& Roth 2008 Dev Cell Anti-phosphotyrosine antibody staining marks the cell outlines (red), DAPI (blue) Index of Other’s Results

  30. Synapsin1Dreyer&Schactner2010 The monoclonal primary antibody from mouse against a fusion protein consisting of a glutathione-S-transferase and the first amino acids of the presynaptic vesicle protein synapsin I coded by its 5′-end (SYNORF1, Klagges et al., 1996 ) Index of Other’s Results

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