immunoYeast & The Antibody Discovery Toolkit Specific - - PowerPoint PPT Presentation

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immunoyeast amp the antibody discovery toolkit specific
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immunoYeast & The Antibody Discovery Toolkit Specific - - PowerPoint PPT Presentation

Apr 11, 2024 839 likes •1.15k views

immunoYeast & The Antibody Discovery Toolkit Specific Antibodies on DemandSo What? Now, you can... Visualize protein localization and identify different cell types: Precipitate proteins to find interaction partners: Measure

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immunoYeast & The Antibody Discovery Toolkit

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Specific Antibodies on Demand…So What? Now, you can...

  • Visualize protein localization and identify different cell types:
  • Precipitate proteins to find interaction partners:

And so much more!

  • Measure specific protein levels:

immunoYeast & The Antibody Discovery Toolkit

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Current State of Technology

From Discovery to Production

immunoYeast Technology

immunoYeast & The Antibody Discovery Toolkit

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Make your own antibodies with immunoYeast!

1.

Obtain diversified antibody library

immunoYeast & The Antibody Discovery Toolkit

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Error-Prone PCR

Antibody Antibody Antibody Antibody

immunoYeast & The Antibody Discovery Toolkit

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Make your own antibodies with immunoYeast!

1.

Obtain diversified antibody library

2.

Select for antibody that binds your POI

immunoYeast & The Antibody Discovery Toolkit

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Error-Prone PCR

Antigen Antigen Antigen Antibody Antibody Antibody

immunoYeast & The Antibody Discovery Toolkit

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immunoYeast & The Antibody Discovery Toolkit

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immunoYeast & The Antibody Discovery Toolkit

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immunoYeast & The Antibody Discovery Toolkit

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Make your own antibodies with immunoYeast!

1.

Obtain diversified antibody library

2.

Select for antibody that binds your POI

3.

Secrete that antibody quickly and easily

immunoYeast & The Antibody Discovery Toolkit

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immunoYeast & The Antibody Discovery Toolkit

Gal80 ¡ Gal4 ¡

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immunoYeast & The Antibody Discovery Toolkit

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immunoYeast The Antibody Discovery Toolkit

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immunoYeast DNA Construction Progress

immunoYeast & The Antibody Discovery Toolkit

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How did we make our constructs?

Biobrick 3A Assembly Overlap Assembly

immunoYeast & The Antibody Discovery Toolkit

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Overlap Assembly

Need to PCR overlapping constructs You can use our overlap oligo script!

immunoYeast & The Antibody Discovery Toolkit

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Overlap Oligo Designer

Available through NYU’s iGEM Wiki

immunoYeast & The Antibody Discovery Toolkit

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Overlap Oligo Designer

immunoYeast & The Antibody Discovery Toolkit

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to iGEM is to Learn

As well as becoming cloning pros.. We learned how to Program! Perl Processing

immunoYeast & The Antibody Discovery Toolkit

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to iGEM is to Learn

and manipulate 3D Structures!

immunoYeast & The Antibody Discovery Toolkit

On the desktop…

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to iGEM is to Learn

…And in the Weill Cornell interactive CAVE!

immunoYeast & The Antibody Discovery Toolkit

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Biobricks in Registry HA Tag C-Myc Tag Biobricks We Made Aga2 Aga2:linker

Current State of Technology

High-throughput in vivo Antibody Discovery K.D. Wittrup, MIT Adimab Tanaka, Rabbitts, Lobato Leeds University, UK

Biobricked Technology

Courtesy of iGEM Osaka

immunoYeast & The Antibody Discovery Toolkit

Antibody Fragment

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LexBD

Biobricked Technology

High-throughput in vivo Antibody Discovery Biobricks in Registry DNA binding domains Transcriptional Activators Biobricks We Made Ura3 Selective Reporter VP16 VP16 LexBD No Interaction Interaction Reporter Gene Reporter Gene LexO LexO

immunoYeast & The Antibody Discovery Toolkit

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Biobricks We Biobricked

Ura3 Selective Marker Aga2 surface display protein Aga2 : linker Secretion Tag

Ab Discovery Toolkit Other Parts:

Suite of yeast promoters w/ RBS Flexible (Gly3)Ser4 linker 36 bp loxP site

immunoYeast & The Antibody Discovery Toolkit

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We couldn’t have done it without:

Gibson, D., Young L., Chuang, R., Venter, J.C., Hutchison, C., Smith, H. Enzymatic assembly of DNA molecules up to several hundred kilobases. Nature Methods 6: 343-345 Chao, G., Lau, W.L., Hackel, B.J., Sazinsky, S.L., Lippow, S.M., Wittrup, K.D. Isolating and Engineering human antibodies using yeast surface display. Nature Protocols 1:755 – 768, 2006. Tanaka, T; Rabbitts T H. Protocol for the selection of single-domain antibody fragments by third generation intracellular antibody capture. Nature Protocols 2: 67-92, 2010. Sauer B. Functional expression of the cre-lox site-specific recombination system in the yeast Saccharomyces cerevisiae. Mol Cell Biol. 7: 2087-96, 1987. immunoYeast & The Antibody Discovery Toolkit

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NYU

  • Dr. David Gresham

Russell Durrett John Phair Harris Kaplan Sung Won Lim Weill Cornell Medical School

  • Dr. Chris Mason

Yossi Steinberger

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Thank You!

immunoYeast & The Antibody Discovery Toolkit