Vivapure adenopack 20 rt

Manufactured by Sartorius

The Vivapure Adenopack 20 RT is a lab equipment product designed for the rapid extraction and purification of adenosine triphosphate (ATP) from various biological samples. It provides a convenient and efficient method for isolating ATP for downstream applications.

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Lab products found in correlation

Bj5183 cells, by Agilent Technologies (2 mentions) Lipofectamine 2000, by Thermo Fisher Scientific (2 mentions) Geneart, by Thermo Fisher Scientific (1 mentions) Gibson assembly, by New England Biolabs (1 mentions) P tetocmv bghpa, by New England Biolabs (1 mentions) Ecorv, by New England Biolabs (1 mentions)

Close spelling variants of this product

Vivapure AdenoPack 20 (8 citations) Vivapure Adenopack (3 citations)

3 protocols using vivapure adenopack 20 rt

Adeno Viral Vector Generation

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To generate the adeno viral vectors (Ad), the selected candidate neoantigens were joined head to tail to generate artificial poly-epitope proteins. A signal peptide was added at N-Term, corresponding to aa 1–29 of the human TPA (tissue plasminogen activator) protein (NP_000921.1). The resulting transgenes were synthesized by GeneART (Thermo Fisher Scientifics) and then transferred into the genome of a Gorilla Adenoviral vector (serotype group C) deleted in E1, E3, and E4 regions and carrying Ad5 E4 ORF6. The resulting recombinant vectors were produced by transfection of adherent M9 cells and amplification in suspension M9 cells. Vectors were then purified from infected cells by Vivapure Adenopack 20 RT (Sartorius).

Leoni G., D’Alise A.M., Tucci F.G., Micarelli E., Garzia I., De Lucia M., Langone F., Nocchi L., Cotugno G., Bartolomeo R., Romano G., Allocca S., Troise F., Nicosia A., Lahm A, & Scarselli E. (2021). VENUS, a Novel Selection Approach to Improve the Accuracy of Neoantigens’ Prediction. Vaccines, 9(8), 880.

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Adenoviral Vector Generation Protocol

Cited 1 time

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Adenoviral vectors were generated as previously described (12 (link)). Briefly, the coding sequences (CDS) for the transgenes encoded in Adenoviral vectors (the corresponding amino acidic sequences are listed in Supplementary Table S1) were synthesized as phosphorylated gBlock dsDNA fragments (IDT). HA tags were added at the N- and C terminus of each transgene. The CDS for all the constructs were generated by Gibson assembly (New England Biolabs) and cloned into the respective shuttle plasmid containing the CMV promoter with two TetO operator repeats and a BGH polyA. The expression cassettes were then transferred into pGAd plasmid, containing the E1/E3/E4 deleted in which the E4 is replaced with Ad5 E4 ORF6. The transgene cassettes were introduced in the E1 deletion locus of related pAdeno by homologous recombination in BJ5183 cells (Agilent). GAd vectors were then produced by transfection of adherent M9 cells (293 cells derivative) with Lipofectamine 2000 (Invitrogen) and amplification in suspension M9 cells. Vectors were then purified from infected cells by Vivapure Adenopack 20 RT (Sartorius). The titer of each vector was determined by qPCR and expressed as viral particles (vp) per mL.

Garzia I., Nocchi L., Avalle L., Troise F., Leoni G., Seclì L., Antonucci L., Cotugno G., Allocca S., Romano G., Conti L., Caiazza C., Mallardo M., Poli V., Scarselli E, & D'Alise A.M. (2024). Tumor Burden Dictates the Neoantigen Features Required to Generate an Effective Cancer Vaccine. Cancer Immunology Research, 12(4), 440-452.

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Recombinant Adenoviral Vector Production

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The coding sequences for CT26-5 and MC38-7 transgenes were purchased as phosphorylated gBlock dsDNA fragments (IDT) and cloned into p-tetOCMV-BGHpA, containg CMV promoter with two TetO repeats and a BGH polyA, previously digested with EcoRV (New England Biolabs). The CDS for CT26-31 was generated by Gibson assembly (New England Biolabs) of two overlapping gBlock sequences (Integrated DNA Technologies) into p-tetOCMV-BGHpA previously digested with EcoRV and Not1 restriction enzymes (New England Biolabs). The expression cassettes were then transferred into pGAd plasmid, containing the E1/E3/E4 deleted in which the E4 is replaced with Ad5 E4 ORF6 of a Great Ape Adenovirus (serotype group C). The transgene cassette was introduced into the E1 deletion by homologous recombination in BJ5183 cells (Agilent). GAds vectors were then produced by transfection of adherent M9 cells (293 cells derivative) with Lipofectamine 2000 (Invitrogen, Thermo Fisher Scientific) and amplification in suspension M9 cells. Vectors were then purified from infected cells by Vivapure Adenopack 20 RT (Sartorius).

D’Alise A.M., Leoni G., Cotugno G., Troise F., Langone F., Fichera I., De Lucia M., Avalle L., Vitale R., Leuzzi A., Bignone V., Di Matteo E., Tucci F.G., Poli V., Lahm A., Catanese M.T., Folgori A., Colloca S., Nicosia A, & Scarselli E. (2019). Adenoviral vaccine targeting multiple neoantigens as strategy to eradicate large tumors combined with checkpoint blockade. Nature Communications, 10, 2688.

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