# Presentation: 423828

## BCR Diagnostics

***Bacterial Chain Reaction for Biomolecular Amplification***

***Jamestown, Rhode Island - bcrbiotech.com***

***Bacteriological Biosensor for Rapid Screening of Platelets Shortly Before Transfusion***

- Boris Rotman, Ph. D.
- Vice President and CSO

## Technical Rationale

**Biosensor integrates three technologies:**
**1.   LEXSAS****TM**** (Label-free Exponential Signal-Amplification System**)
- 	    A recently developed system using engineered spores as fluorogenic nanodetectors.
**2.**	**Microfluidics: **
-            The detection system is a unique device containing an array of 80,000 **microscopic, independent** biosensors, each of which is about 5-picoliter volume. This small reaction volume enables detection of **individual** bacterial cells. The device is  termed “80K-bioChip.”
**3.   Computerized Parallel Imaging**
- 	    The 80K-bioChip provides high-throughput data acquisition and processing.

## LEXSASTM Characteristics

**Ability to detect and count low bacterial levels.**
**Quantitative results in less than 20 minutes.**
**Cost-effective testing. **
**Linear detection response over a wide dynamic range of one to 10,000 bacteria per sample.**
**Low instrumentation cost.**
**Applicability to automated high-throughput operations.**

## Why using Engineered Spores for Cell-based Biosensing? 

**Spores are ****metabolically dormant**** microorganisms resistant to extreme chemical and physical environmental conditions. **
**Despite their ruggedness, however, spores ****remain alert**** to specific environmental signals that ****trigger the spores**** to rapidly break dormancy and initiate germination.**
**The spores in the LEXSASTM** **are ****engineered to be fluorogenic****, i.e., the spores are ****not**** fluorescent per se,  but produce fluorescent light in the presence of nearby bacteria. **

## Principle of the Biosensor

_***Germination Signals***_

*** ***

_*** ***__***Analyte + germinogenic buffer***_

**SPORE**

**IMAGE ANALYSIS**

_***Signal Receptor***_

**DATA PROCESSING AND DISPLAY**

_**Transduced Fluorescent Signals**_

_***Spores Engineered***_
***as***
_***Fluorogenic Nanodetectors ***_

**Bacteria**

## Response of Engineered Spores to Germinants

## Schematic of 80K-bioChip[Cross Section]

## SEM image of 80K-bioChip[Micro-colanders® are 20 μm dia]

## SEM image of Micro-colander®  with about 200 Spores

## 80K-bioChip mounted on filtration cartridge

## Micro-colander® Scanner

## Biosensor Operation

**Platelet sample is processed to separate bacteria, and the**
**resulting suspension is mixed with engineered spores**
** **
**Mixture is combined with germinogenic substrate and then filtered through the 80K-bioChip**
**80K-bioChip is incubated at 37****o****C**
**Fluorescent images of 80K-bioChip are captured at time intervals**
**Digital  images are processed to yield number**** of bacteria in the sample **

## Schematic of 80K-bioChip[Top View]

**Micro-colanders with one bacterium show visible fluorescence**

## 80K-BioChip showing ten (10) micro-colanders with significant fluorescence above baseline

**80K-BioChip showing ten (10) micro-colanders with significant fluorescence above baseline**

## Photomicrographs of 80K-bioChip

- Left:  Normal light                Right:  Fluorescence light

## BCR Diagnostics, Inc.

- Privately held since 1994
- Technology protected by five U. S. and multiple worldwide Patents  (additional U.S. and ROW pending)
- Awarded Phase I and Phase II Grants from the    Rhode Island Center for Cellular Medicine
- Awarded Phase I and Phase II SBIR Grants from the National Institutes of Health

## Publications

- Rotman, B., Cote, M. A., and Ferencko, L.  Phenotypic Engineering of Spores. U. S. Patent pending.
- Rotman, B.  Analytical system based upon spore germination. U. S. Patent No.6,596,496, March 29, 2005.*
- Ferencko, L., Cote, M. A., and Rotman, B.  Esterase activity as a novel parameter of spore germination.  Biochem. Biophys. Res. Comm., _319_: 854-858, 2004.
- Rotman, B. and Cote, M. A.  Application of a real-time biosensor to detect bacteria in platelet concentrates.  Biochem. Biophys. Res. Comm., _300_, 197-200, 2003.
- Rotman, B.  Using living spores for real-time biosensing.  Gen. Eng. News, _21_, 65, 2001.
- *Additional patents listed at www.bcrbiotech.com