Revolutionizing Life Science Moxi Flow ™ Increase efficiency and accuracy in yeast counting FELC 2014

Revolutionizing Life Science ACCURATE Gold standard precision and accuracy 30K events measured in 10 seconds Integrated Coulter Principle counts Eliminates user-to-user variability Propidium iodide is 20-50X more sensitive than methylene blue exclusion EASY TO USE No cleaning or maintenance Automatic bud count capability Requires minimal training, intuitive smart phone interface Can store gates for up to 5 yeast strains FAST Counts yeast in less than 10 seconds Moxi Flow Yeast Counter

Revolutionizing Life Science Confidential, not to be shared outside of Orflo or it's Distributors Accurate yeast counts are critical to the success of the propagation and fermentation process Hemocytometers are used by 95% of the fuel ethanol industry Yeast counts are done 5-10 times per day To ensure accuracy, hemocytometer counts are done in triplicate

Revolutionizing Life Science Confidential, not to be shared outside of Orflo or it's Distributors Time Consuming Poor Statistical Robustness User-to- User Variability 5-10 counts per day Increase labor cost Counts need to be done in triplicate Increase labor cost Inaccurate counts Wasted material Longer process Low yield Risk entire ferm

Revolutionizing Life Science Confidential, not to be shared outside of Orflo or it's Distributors Accurate Yeast Counts Use material and labor more efficiently Increase in ethanol yield Decrease lag time in the fermentor Identify yeast in log phase Identify problems earlier

Revolutionizing Life Science Confidential, not to be shared outside of Orflo or it's Distributors Select Yeast Count assay by touching desired button

Revolutionizing Life Science Confidential, not to be shared outside of Orflo or it's Distributors Step by step instructions given on screen Choose the size specific to your yeast strain

Revolutionizing Life Science Confidential, not to be shared outside of Orflo or it's Distributors 1. Open top and bottom doors of unit 3. Close both doors 2. Insert the cassette

Revolutionizing Life Science Confidential, not to be shared outside of Orflo or it's Distributors 1. Open top door only 3.Close doors 4.The test will begin running and plot data in real-time 2.Rapidly pipette 75ul sample into well of cassette Do not worry about a bead forming

Revolutionizing Life Science Confidential, not to be shared outside of Orflo or it's Distributors Note this is the valley between budding yeast (to the right) and single cell yeast population (to the left) Total % Viability is listed in the yellow box at the top of the screen (77.9%)

Revolutionizing Life Science Confidential, not to be shared outside of Orflo or it's Distributors Note this is the valley between budding yeast (to the right) and single cell yeast population (to the left % Budding = 3.98/6.21 = 64% Press RCL button and select “1” Record live budding cell count (3.98 x 10^4) Press RCL button and select “1” Record live budding cell count (3.98 x 10^4)

Revolutionizing Life Science Confidential Notes: Highly repeatable counts, typical CV’s less than 5% Within 5% of hemocytometer despite thickness of starting slurry Four Repeats

Revolutionizing Life Science Confidential Fig. 4– Average coefficients of variation (CV) percentages for both counts and viability measurements for all systems.

Revolutionizing Life Science Confidential Get accurate counts in less than 10 seconds Reduce user error Decrease lag time in fermentor Increase Efficiency Discover trends in the propagation and fermentation process Identify bacterial contamination Identify Trends Propagate with healthy budding yeast Get a better grasp on the variables that help maximize ethanol production Maximize Yield

Revolutionizing Life Science Confidential