Leukemia Components of the bone marrow Eosinophil Myelocyte - - PowerPoint PPT Presentation

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Leukemia Components of the bone marrow Eosinophil Myelocyte - - PowerPoint PPT Presentation

Mar 09, 2024 408 likes •772 views

Leukemia Every 4 minutes a new person is diagnosed Every 10 minutes somebody dies 10% of all leukemia patients are under 20 Team 2010 BIOTEC Dresden Svea & Mareike ricks ensor iology ynthetic Leukemia Components of the bone marrow

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Every 4 minutes a new person is diagnosed Every 10 minutes somebody dies

10% of all leukemia patients are under 20

Leukemia

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ensor

ricks

Team 2010 BIOTEC Dresden

Svea & Mareike

iology ynthetic

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Leukemia

Myelocyte Erythrocyte Normoblast Fat Eosinophil

Components of the bone marrow

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Current Leukemia Diagnosis

Anti-CD33 CD33+ cell

State of the art

FACS

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Desired Features of SensorBricks

Simplicity Modularity Sensitivity

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SensorBricks - ToolBox

Domain B of Protein A

Protein A from Staphyloccocus aureus

CD33+ cell

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SensorBricks - ToolBox

SAM AHL LuxI LuxR

Quorum Sensing System from Vibrio fischeri

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SensorBricks - ToolBox

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patients blood cells Mix anti-CD33 + Fusionprotein

SensorBricks in Real Life

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patients blood cells Mix anti-CD33 + Fusionprotein Antibody- Fusionprotein + blood sample

SensorBricks in Real Life

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patients blood cells Mix anti-CD33 + Fusionprotein Antibody- Fusionprotein + blood sample Take SN containing AHL

SensorBricks in Real Life

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add sensor Detect fluorescent signal

SensorBricks in Real Life

pTet LuxR pLuxR YFP patients blood cells Mix anti-CD33 + Fusionprotein Antibody- Fusionprotein + blood sample Take SN containing AHL

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Experimental Pipeline

Build Detection System Test SensorBricks receiver Simulate AHL sender Test SensorBricks sender Establish Leukemia Test

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Results

Build Detection System Test SensorBricks receiver Simulate AHL sender Test SensorBricks sender Establish Leukemia Test

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pTet LuxR pLuxR YFP

Building a Detection System

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pTet RFP pTet LuxR pLuxR YFP

Building a Detection system

Building a Detection System

RFC 78

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pTet RFP pTet LuxR pLuxR YFP

SensorBricks in Real Life

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pTet RFP pTet LuxR pLuxR YFP

SensorBricks in Real Life

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Build Detection System Test SensorBricks receiver Simulate AHL sender Test SensorBricks sender Establish Leukemia Test

Results

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Results

Build Detection System Test SensorBricks receiver Simulate AHL sender Test SensorBricks sender Establish Leukemia Test

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Test SensorBricks Receiver

The fluorescence signal is proportional to the AHL input

12,00 14,00 16,00 18,00 20,00 22,00 24,00 26,00 20 40 60 80 100 120 140

2000 1000 500 200 100 50 10 1 0,1 0,01

Fluorescence (YFP/RFP) Time in minutes [AHL] in nM

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Direct correlation of fluorescence to [AHL] after 2 hours

Fluorescence (YFP/RFP) AHL concentration in nM

10 12 14 16 18 20 22 500 1000 1500 2000

Test SensorBricks Receiver

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Results

Build Detection System Test SensorBricks receiver Simulate AHL sender Test SensorBricks sender Establish Leukemia Test

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Results

Build Detection System Test SensorBricks receiver Simulate AHL sender Test SensorBricks sender Establish Leukemia Test

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pTet LuxI

Receiver detecting AHL Sender expressing LuxI

Simulate AHL Sender

Proof of Principle: Cocultivation Assay

pTet RFP pTet LuxR pLuxR YFP

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20 40 60 80 100 120 140 Sender Receiver +500 nM AHL 20:1 100:1 1000:1 10000:1

Low amounts of LuxI lead to a detectable induction

Ratio Receiver : Sender Fluorescence (YFP/RFP)

Simulate AHL Sender

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Results

Build Detection System Test SensorBricks receiver Simulate AHL sender Test SensorBricks sender Establish Leukemia Test

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Future Outlook

Build Detection System Test SensorBricks receiver Simulate AHL sender Test SensorBricks sender Establish Leukemia Test

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Future Outlook

Build Detection System Test SensorBricks receiver Simulate AHL sender Test SensorBricks sender Establish Leukemia Test

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Potential of SensorBricks

Amplification

pTet RFP pTet LuxR pLuxR LuxI YFP

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Potential of SensorBricks

Amplification Pigments

http://2009.igem.org/Team:Cambridge

pTet RFP pTet LuxR pLuxR LuxI YFP
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Potential of SensorBricks

Amplification Modularity Multiple Detection

pTet RFP pTet LuxR pLuxR LuxI YFP

Pigments

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Thanks to: Andy Oates, Annelie Oswald, Marko Storch, Petra Schwille, Johnson Madrid, Kaj Bernhardt, Michael Bachmann BIOTEC Dresden, MPI-CBG Dresden, TU Dresden

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FACS Facts

Feature FACS SensorBricks Detection Limit 1 cell in 10 000 cells 1 sender cell in 10 000 receiver cells Price (Hardware) in US$ ~ 200 000 ~ 50 000 Total test time 3 h 4-5 h