The antibodies to BLV were detected using a commercially available verification ELISA kit (Pourquier, Montpellier, France) following the manufacturer's instructions.
After sequence verification (Big Dye kit, ABI Perking Elmer) the pET28-1738 plasmid was transformed into E. coli BL21 (DE3) selecting for resistance to kanamycin (50 μg.ml-1).
Validation included manufacturing of 1 1 parts and mockups, test of prototypes in the FLEX and HELOKA-LP helium loops of KIT for verification of the thermal and mechanical properties, and irradiation of specimen filled capsule prototypes in the BR2 test reactor.
Though specificity testing was not a goal of this paper, the kit manufacturer's verification documentation states that 100% of 40 strains of non-target organisms including related genera (e.g., Bacillus spp., Listeria spp).
After DNA purification using the Qiagen Midi Plasmid Kit (Qiagen) and verification of the inserted 5 kb fragment by sequencing (Eurofins MWG Operon), HaCaT cells were transfected with the indicated reporter plasmid and the empty pGL4.74 renilla control vector (Promega).
The ends of the inserted PCR product were sequenced using the primers provided with the cloning kit for additional verification of FIV Ple cloned products.
If, at any stage, the test procedure associated with the performance specification is modified (eg, if reaction volumes of a CE-marked kit are reduced), verification is not appropriate and validation is required.
Furthermore, together with specific staining after trilineage differentiation, the use of commercially available functional kits provides a systematic verification of MSC identity, independently of the tissue or species type.
Sporadic MSI CRCs were additionally studied with the SALSA MS-MLPA ME042-B2 CIMP kit (MRC-Holland, Amsterdam, The Netherlands) for verification.
