Table 1.
TMEFF2 immunoblot studies.
| Cells | Reducing/Non-Reducing | Endogenous/Overexpressed | Antibody Used | Band Sizes kDa | Detail | Ref. |
|---|---|---|---|---|---|---|
| N2a | Not mentioned | Overexpressed | Anti-V5 | 17, 50, 56 | Duplet band between 50 and 56 kDa, plus a single band just below 17 kDa (C-terminal band) | [7] |
| CWR22 | Not mentioned | Endogenous | 2H-8 a | 51, 58 | Duplet band at 51–58 kDa | [18] |
| LNCap, C4-2 | Not mentioned | Endogenous | TMEFF-CP b | 44 | Single 44 kDa band in both LNCap and C4-2 cells | [19] |
| 22Rv1, LNCap | Not mentioned | Endogenous | Abcam | ~ 48 | Single band just under 50 kDa marker (size not mentioned in text) | [20] |
| 22Rv1, LNCap | Reducing | Endogenous | Abcam | 40 to 55 | Single smear ranging from 40 to 55 kDa in LNCap and doublet band between 40 and 55 kDa in 22Rv1 cells | [21] |
| RWPE1, RWPE2 | Not mentioned | Overexpressed | Abcam | 40 to 56 | A smear from 40–56 kDa in RWPE1 and RWPE2 cells, a heavily glycosylated band can be observed above 90 kDa in only the TMEFF2_ΔGA form | [22] |
| HEK293 | Reducing | Overexpressed | 11D1C1 c | 58 | Full length at 58 kDa, ECD only at 54 kDa (bands appear as duplets) | [23] |
| PC3, LNCap | Non-reducing | Both (see detail) | 2H-8 | 43, 48, 52, 97 | Endogenous TMEFF2 was detected at a single band of 43 kDa in LNCaP cells. V5/His tagged TMEFF2 overexpressing PC3 cells showed a major band at 48 kDa, a minor one at approximately 52kDa and another (smear) band at approximately 97 kDa. ECD-only overexpressing PC3 cells showed a band at 40 kDa. | [15] |
| CHO | Reducing | Overexpressed | Anti-V5 | 14, 71 | V5/His tagged overexpressing TMEFF2 in CHO cells showed two distinct bands: one at 71 kDa and another at 14 kDa. For ECD-only overexpressing CHO cells, a duplet band at 54–60 was detected in cell lysate while 4 bands were detected in the medium, at 99, 69, 63 and 52 kDa. | [12] |
| CHO, A172 | Both (see detail) | Both (see detail) | 2H-8, aTR-C d | 51 | In A172 cells, endogenous TRc was detected at 51 kDa under non-reducing conditions (2H8 antibody). For TRc-overexpressing CHO cells, the band was observed at 51 kDa under non-reducing conditions and at 58 kDa under reducing conditions in cell lysates (pAb-aTRc antibody). In the media of TRc-overexpressing CHO cells, under non-reducing conditions, a smear was observed at 39–45 kDa (2H8 antibody). | [11] |
| HEK293, LNCaP, CHO, PC3 | Not mentioned | Both (see detail) | Anti-V5, Anti-TMEFF2 (details not mentioned) | 60, 75, 10, 14, 22 | Endogenous TMEFF2 from LNCap cells, the shed form from media, showed a duplet band at 60–75 kDa. In TMEFF2/V5/His overexpressing HEK293, a duplet band at 60–75 kDa was detected in conditioned media (atni-TMEFF2 antibody) and 10, 14 and 22 kDa distinct bands were detected in the cell lysate (anti-V5 antibody). Similarly, in TMEFF2/V5/His overexpressing CHO and PC3 cells, 10, 14 and 22 kDa c-terminal bands were detected in the cell lysate using V5 antibody. | [5] |
| HEK293 | Not mentioned | Overexpressed | Anti-V5 | 15, 17, 20, 24, 28 | Only C terminal bands were mentioned. In HEK293 cells stably transfected with AP-TMEFF2-V5/His, bands changed upon overexpression of various proteinases: (Matriptase-1) 24 and 28 kDa, and (Hepsin) 15 and 20 kDa. A 17-kDa band was observed without overexpression of any protease and was proposed to be a result of native ADAM activity. | [17] |
a 2H-8 mAb was generated against TR extracellular domain [11]. b TMEFF-CP pAb was generated against a.a 345–359 of TMEFF2 cytoplasmic domain [19]. c 11D1C1 mAb was generated against TMEFF2 extracellular domain by DNA immunization [23]. d aTR-C pAb was generated against a.a. 346–360 of TRc cytoplasmic domain [11].