Determination of Mammalian Cell Counts, Cell Size and Cell Health Using the Moxi Z Mini Automated Cell Counter

The Mxi Z Mini automated cell counter is an ultra small benchtop instrument that performs highly accurate cell and particle counts. By combining the gold standard TER principle with a patented thin film sensor technology. The cassettes used with the Mxi Z are precision microfluidic sensors to obtain cell counts.

A cassette is inserted in the front of the unit and 75 microliters of sample is loaded into the sample port. When the run begins, cells are pulled single file through a cell sensing zone in the thin film membrane of the cassette, thousands of cells per sample accounted as they travel through the sensing zone. The precise volume of each is determined by an impedance measurement.

After eight seconds, the cell concentration average cell size and cell health index are displayed along with a complete high resolution histogram of the whole particle population. In the sample, the sample data can then be analyzed and stored on the instrument itself or transferred to a Mac or PC via Bluetooth or USB. The main advantage of this technique over other systems such as hemo, cytometers, or image-based automated cell counters is that it is far more reliable with accuracy and precision greater than 95%as compared to 75 to 80%with image-based systems.

Plus it measures cell size and it is much faster and it is not subjective. Though this method can is ideal from a million cell counts. It can be applied to a wide variety of cell counting applications because of its broad dynamic range.

Such applications include white blood cell counts, red blood cells, protozoa, algae, and yeast. Begin this experiment by diluting a freshly detached suspension of HEK 2 93 cells in a medium that has physiological conductivity, such as all flow diluent or PBS to the appropriate concentration. Gently triturate to minimize cell clustering, ensure that the unit is unplugged from the power supply.

Then turn the mxi Z on By pressing the power button, the home screen will be displayed. The Mxi Z uses disposable type M or S cassettes, which are precision microfluidic sensors. Each cassette has two loading ports indicated by blue oval and labeled one or two.

Each cassette can be used for two tests. Type M cassettes have an operating range of 3000 to 500, 000 cells per milliliter. The range for type S cassettes is 3000 to 2.5 million cells per milliliter.

Press the tray down and insert a cassette to ensure homogeneity of the sample slowly inverted several times. Then pipette 75 microliters of sample. Place the pipette tip firmly into the sample port, holding the pipette at a 45 degree angle, and then dispense the sample into the port in one smooth motion, ensuring the tall of the sample has been dispensed.

Touch the screen in the designated circle or anywhere except in the small particle mode box to start counting. The mxi Z will begin the test and the cell count results will be complete in eight seconds. With the Type M cassette, the results are automatically displayed with a proprietary curve fitting algorithm that fits the single cell population of interest and also corrects the total count for coincidence.

Events or events where multiple particles or clusters pass through the cell sensing zone simultaneously. Note that while these coincidence events fall outside the curve fit area to the right, they're still included in the total counts. Once a suitable fit is found, a high resolution histogram of the entire sample population is displayed along with the numerical values for cell concentration, average diameter, and average volume.

In addition, a moxie population index or MPI value is given as an indicator of general culture health. The counts at the far left of the histogram are normal and represent the baseline noise threshold of the system adjusted to optimize system sensitivity and possible sample debris counts. Once the sample run is complete, remove the cassette from the unit select done to save and return to the home screen.

A visual indicator confirms the sample has been run. If a second sample is to be run, turn the cassette around and insert it into the un. Again, following a run in the normal mode, the results are displayed for a curve fit count on a size scale of four to 34 microns for the type M cassette or three to 26 microns for type S to rescale the x axis to four to 26 microns, touch the rescale icon.

Once successively, press the rescale icon to change the x-axis scale through the following scales, four to 18 microns, four to 10 microns, and back to four to 34 microns. Toggle back to curve fit mode if desired. The test number for the current analysis is displayed at the top of the histogram in a green box.

The data will be saved as the test number touch the gating icon to toggle the gating mode, which allows for manual gating of the histogram. This makes gating the default acquisition mode. Gates may be changed by sliding the left and right blue gating markers or by touching either marker and then incrementally moving them with the arrow keys that appear.

The displayed counts and average sizing information are calculated only from the particles within the gating range. Press the done icon to save the results and return to the home screen. To open a save test, press the histogram icon on the home screen.

Icons for up to nine saved histograms will be displayed at a time touch page down or page up or swipe display up or down. To view more test results as needed and select the icon for the test of interest. Each histogram will be opened in either the curve fit mode or gated mode.

Depending on how it was saved, the cassette type used for the test, either M or S will be indicated in the top left of the screen.Saved. Histograms can also be toggled between gating and curve fit modes to auto gate touch the desired peak. This sets the optimum gait positions for the selected peak.

This is especially useful for multi peak situations. To indicate a peak of interest, press the zoom and unzoom arrow icons to adjust the count scale on the Y axis of the histogram. The vertical scale can also be changed by swiping a finger over the peak upwards to decrease or downwards to increase press the back icon to close this test and return to the home screen.

Data can be transferred to a Mac or PC using the moxie chart application. To do this first, press the Bluetooth icon on the home screen of the Moxie Z unit. The unit will report its Bluetooth ID on the computer, open the moxie chart application and click on the Bluetooth icon in the upper right corner.

Then choose the device that corresponds to the Bluetooth ID of the Z and selected destination folder for the uploaded files in addition to Bluetooth. Another way to transfer data from the Mxi Z to a computer is by using a USB cable and treating it as a normal USB device. With data files, data files from a Mxi Z unit connected via A USB can be copied to any directory on the computer.

Files can be opened and analyzed directly with the Mxi Charles application or because they are formatted CSV files, they can be directly opened in Microsoft Excel or other data analysis programs for customer analysis to compare the results of counting in the Mxi Z to those of obtained. Using a cool to counter identical suspensions of precision calibrated beads and mammalian cells were counted by both methods. Four different sized beads ranging from six to 15.6 microns in diameter, plus several different cell lines were tested.

The average of three to four counts was plotted as can be seen here. The counts obtained were nearly identical, indicating that the Moxie Z is capable of obtaining accurate counts in a wide range of sizes to demonstrate the range of concentrations that can be accurately assessed using the Mxi Z.The same beads and cells were serially diluted from an initial 500, 000 cells per milliliter for the type M cassette or roughly 2.5 million cells per milliliter for the type S.The theoretical cell concentrations were then plotted against the counts obtained as shown here. The measured concentrations matched the theoretical cell concentrations demonstrating accurate counting across the dynamic range of each cassette, 3000 to 500, 000 cells per milliliter for the type M cassettes and 3000 to 2.5 million cells per milliliter for type S ca to examine the variability of the results obtained five separate counts of HEK 2 93 and HELOC cells in the 200, 000 to 300, 000 cells per milliliter range were obtained using a call to Z two, Moxi Z and hemo cytometer.

As can be seen in this graph, the mean coefficient of variation was comparable in the Z two and mxi Z and much less than that of the hemo cytometer indicating that the Mxi Z's reproducibility is comparable to that of a call to Z two and superior to that of a Hema cytometer. To demonstrate the precision of the Mxi Z in determining particle size, manufactured beads were measured using the Mxi Z five Measurements for each bead size were measured, averaged, and plotted against the manufacturer reported bead size. The Mxi Z measured the beads at the size reported by the manufacturer.

The R squared value was 0.9989 to demonstrate the ability of the mxi Z to assess culture. Health cell health index measurements were recorded using the Mxi Z for identical dead cell spike, HEK and hela cell preparations, and compared to the visual trian blue exclusion results of the same sample hemotoma viability percent or flow cytometer guava PCA for comparison purposes. Viability measurements for the samples were also plotted on the graph as can be seen here.

The calculated viability for both HGK and LAR cells obtained using the Mxi Z were on par with the viability measurements obtained using the Trian blue exclusion method with r squared values of 0.9937 and 0.9728 respectively. The data obtained are also comparable to that obtained using guava PCA with the flow cytometer with R squared values of 0.9724 for he EK cells and 0.9585 the hela cells. While in certain cases the cell health index may agree with standard viability approaches, it is an entirely new and different technique to health assessment and consequently or not always be identical to the older techniques.

After watching this video, you should have a good understanding of how to easily obtain cell health, cell size and cell count information While attempting this procedure. It is important to make sure that the cell density is in the correct range.