Trop-2 and Integrin Immunoprecipitation and Immunoblotting

The T16 mouse monoclonal antibody (mAb) against Trop-2 (gift of Dr. S. Alberti) and the TS2/16 mAb against β1 integrin (HB-243, ATCC) were used for IP. The following Abs were used for IB: mAbs against β1 integrin subunit (610468, BD Transduction Laboratories), CD63 (Ab8219, Abcam), CD81 (Ab23505, Abcam); goat polyclonal Abs (pAbs) against human and murine Trop-2 (AF650 and AF1122, R&D Biosystems); rabbit antisera against α3 and αv integrin subunits; rabbit pAbs against α5 integrin subunit (sc-10729, Santa Cruz Biotechnology), β5 integrin subunit (AB1926, Millipore), Calnexin (sc-11397, Santa Cruz Biotechnology) and ERK1/2 (sc-93, Santa Cruz Biotechnology). The goat pAb against human Trop-2 (AF650, R&D Biosystems), a mAb to Vinculin (MAB1624, Millipore) and a mAb to FAK (05-537, Millipore) were used for IF. A goat pAb against human Trop-2 (R&D Biosystems) was used for IHC. Non-immune goat IgG (SantaCruz), non-immune rabbit IgG (Sigma) and non-immune mouse IgG (Pierce) were used as negative control Abs. FN purification from human plasma has been previously described [17 (link)].

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Trerotola M., Ganguly K.K., Fazli L., Fedele C., Lu H., Dutta A., Liu Q., De Angelis T., Riddell L.W., Riobo N.A., Gleave M.E., Zoubeidi A., Pestell R.G., Altieri D.C, & Languino L.R. (2015). Trop-2 is up-regulated in invasive prostate cancer and displaces FAK from focal contacts. Oncotarget, 6(16), 14318-14328.

Publication 2015

HB-243 Ab8219 Ab23505 AF650 sc-10729 sc-11397 sc-93 non-immune rabbit IgG

Antisera Calnexin Erk1 Goat Human Integrin Monoclonal antibody Mouse Plasma Rabbit Subunit Trop 2 Vinculin αv integrin β1 integrin

Corresponding Organization :

Other organizations : University of Chieti-Pescara, Thomas Jefferson University, Sidney Kimmel Cancer Center, University of British Columbia, The Wistar Institute

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Variable analysis

independent variables

T16 mouse monoclonal antibody (mAb) against Trop-2
TS2/16 mAb against β1 integrin

dependent variables

Protein expression levels of β1 integrin subunit, CD63, CD81, α3 integrin subunit, αv integrin subunit, α5 integrin subunit, β5 integrin subunit, Calnexin, ERK1/2

control variables

Non-immune goat IgG
Non-immune rabbit IgG
Non-immune mouse IgG

controls

Positive control: Fibronectin (FN) purified from human plasma
Negative controls: Non-immune goat IgG, non-immune rabbit IgG, non-immune mouse IgG

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