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esp3i enzyme  (Twist Bioscience)
90
Twist Bioscience esp3i enzyme
Novel monoclonal antibodies were generated via mouse immunization and hybridoma screening. Four monoclonal antibodies are shown which have high reactivity to (A) Jurkat T cells and (B) primary human T cells. Following this, antibody heavy and light chains were sequenced. (C) scFv DNA was then synthesized and assembled into CD19 or EGFRvIII BiTE molecules using single-pot restriction ligation with <t>Esp3I</t> restriction enzyme. (D) Ten unique CD19 or EGFRvIII BiTE plasmids were then transfected into HEK293T cells and supernatants were tested using Jurkat cells in 1:1 co-cultures with EGFR-vIII+ U87vIII cells ( left ) or CD19+ Raji cells ( right ) using varying doses of BiTE supernatant as shown. (E) Four BiTE plasmids found to produce active BiTE molecules were tested again against EGFRvIII+ U87-vIII cells, EGFRvIII- U87-WT cells, or CD19+ Raji cells. (F) BiTE supernatants for CD19 or EGFRvIII targeted molecules incorporating the 1E2 CD3-scFv were added to co-cultures with 10 000 primary human T cells and 2000 target cells. Graphs show the relative fluorescent signal of red-fluorescent target cells over 5 days in co-culture. Results are derived from a single experiment, but are representative of at least 3 repeated observations.
Esp3i Enzyme, supplied by Twist Bioscience, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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esp3i enzyme - by Bioz Stars, 2026-03
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Novel monoclonal antibodies were generated via mouse immunization and hybridoma screening. Four monoclonal antibodies are shown which have high reactivity to (A) Jurkat T cells and (B) primary human T cells. Following this, antibody heavy and light chains were sequenced. (C) scFv DNA was then synthesized and assembled into CD19 or EGFRvIII BiTE molecules using single-pot restriction ligation with Esp3I restriction enzyme. (D) Ten unique CD19 or EGFRvIII BiTE plasmids were then transfected into HEK293T cells and supernatants were tested using Jurkat cells in 1:1 co-cultures with EGFR-vIII+ U87vIII cells ( left ) or CD19+ Raji cells ( right ) using varying doses of BiTE supernatant as shown. (E) Four BiTE plasmids found to produce active BiTE molecules were tested again against EGFRvIII+ U87-vIII cells, EGFRvIII- U87-WT cells, or CD19+ Raji cells. (F) BiTE supernatants for CD19 or EGFRvIII targeted molecules incorporating the 1E2 CD3-scFv were added to co-cultures with 10 000 primary human T cells and 2000 target cells. Graphs show the relative fluorescent signal of red-fluorescent target cells over 5 days in co-culture. Results are derived from a single experiment, but are representative of at least 3 repeated observations.

Journal: PLOS ONE

Article Title: A simplified function-first method for the discovery and optimization of bispecific immune engaging antibodies

doi: 10.1371/journal.pone.0273884

Figure Lengend Snippet: Novel monoclonal antibodies were generated via mouse immunization and hybridoma screening. Four monoclonal antibodies are shown which have high reactivity to (A) Jurkat T cells and (B) primary human T cells. Following this, antibody heavy and light chains were sequenced. (C) scFv DNA was then synthesized and assembled into CD19 or EGFRvIII BiTE molecules using single-pot restriction ligation with Esp3I restriction enzyme. (D) Ten unique CD19 or EGFRvIII BiTE plasmids were then transfected into HEK293T cells and supernatants were tested using Jurkat cells in 1:1 co-cultures with EGFR-vIII+ U87vIII cells ( left ) or CD19+ Raji cells ( right ) using varying doses of BiTE supernatant as shown. (E) Four BiTE plasmids found to produce active BiTE molecules were tested again against EGFRvIII+ U87-vIII cells, EGFRvIII- U87-WT cells, or CD19+ Raji cells. (F) BiTE supernatants for CD19 or EGFRvIII targeted molecules incorporating the 1E2 CD3-scFv were added to co-cultures with 10 000 primary human T cells and 2000 target cells. Graphs show the relative fluorescent signal of red-fluorescent target cells over 5 days in co-culture. Results are derived from a single experiment, but are representative of at least 3 repeated observations.

Article Snippet: Swapping CD3-targeting domains was performed similarly as described above using Esp3I enzyme to integrate synthetic CD3-scFv fragments (Twist Biosciences, USA), derived from in-house generated CD3-specific murine monoclonal antibodies, into the pBiTE construct.

Techniques: Bioprocessing, Generated, Synthesized, Ligation, Transfection, Co-Culture Assay, Derivative Assay