PRECISION DEVICE FOR THE RAPID DIAGNOSTIC OF MICROVILLUS INCLUSION DISEASE Theodore J Hall, Biomedical Engineer

Dr. Michael Caplan Ph.D. , School of Biological Health Systems and Engineering David Carpentieri, M.D. , Mitchell Shub, M.D. , Phoenix Children’s Hospital

-Microvillus Inclusion Disease (MVID) is a rare genetic disease which leads to a non-functioning brush border in the small intestine. -Due to the 1977 CàT single nucleotide mutation, this disease un-proportionally affects people of the Navajo nation. -Current diagnosis methods have patients waiting over 4 weeks for results, a critical window for patient survivability. With implementation of the Tentacle Probe (TP), diagnostic cost and time is reduced to $100 and 24 hours.

Background

Equations

Legend pr is the probability of reaction; v is the average velocity R is the sum of the radii of the two molecules; P is the probe concentration T is the target concentration; kf is the association rate constant

[1] Erickson, Robert P., et al. American Journal of Medical Genetics. Part A: Navajo Microvillous Inclusion Disease is due to a Mutation in MYO5B. 146A Vol. Wiley-Liss, 12/15/2008. Web. 22 Nov. 2015. [2] Satterfield, B.C., J.A.a. West, and M.R. Caplan. “Tentacle Probes: Eliminating False Postives without Sacrificing Sensitivity.” Nucleic Acids Research(2007). Print.. [3] Christine Beeson, Jeffrey C. Weiss, Robert D. Ligensky. Microvillus Inclusion Disease- A Deadly Masquerador.

 

[2] Works Cited [2]

[2]

Future Work

Methods -Diagnostic device will follow the steps: 1) DNA acquisition via blood sample 2) DNA purification 3) rt-PCR analysis of DNA with TP 3) output result of patient diagnosis -Development of wild type probe has the ability to predict the likelihood of passing down the mutation to further generations

    Sequence of the TP: (5’  -­‐  [BHQ1]  -­‐  ccgCAAGCGTTGAGCTTGcgg  -­‐  [orange  560]  -­‐  [linker  9]  -­‐

GACATAGTGAGGTGTCGTGGCA  -­‐  3’)  

The TP combined with DNA containing the target mutation was run through a melting curve. As temperature increased the probe and DNA melted, allowing for a binding event to occur.

The binding event demonstrates that the probe will bind to the target mutation

-Tentacle Probe (TP) is used in a rt-PCR machine. When the probe binds to the target mutation it will fluoresce; as shown in the melt curve. -The mutation is detected by the levels of fluorescence. The resulting exponential fluorescence curve serves as the diagnostic determinant. -Since an rt-PCR machine is used to amplify the DNA and the TP is used to detect the mutation, a rapid and accurate diagnostic result can be obtained

Results