Practical Guide to Sony SH800 Sorter

Sony SH800 Flow Cytometer Sorter: Full Process Operation and Maintenance Guide from Startup to Shutdown

Flow cytometry sorting is an indispensable technology in life sciences and biomedical research. The Sony SH800 series cell sorter, with its innovative microfluidic chip design, intuitive software interface, and stable flow system, has become an important tool for many core facilities and laboratories. However, in order to fully utilize its performance and extend the lifespan of the instrument, operators must strictly follow a standardized set of start-up, calibration, experimentation, and shutdown procedures. This guide will be based on the original factory technical manual and provide users with a complete technical document covering system characteristics, daily inspections, key operating steps, and common precautions.

Chapter 1: System Overview and Core Features

The Sony SH800 sorter is a desktop cell sorting system that uses disposable microfluidic chips. Its design significantly reduces the risk of cross contamination and simplifies daily maintenance. Understanding its hardware composition is the foundation for correct operation.

1.1 Laser and Optical Path System

The instrument comes standard with four solid-state lasers, with wavelengths of:

405 nm

488 nm

561 nm

638 nm

It should be noted that the laser spot is of a single size (1), which means that strict compensation adjustments must be made when using fluorescent dyes with overlapping emission spectra, otherwise it will lead to data parsing errors.

1.2 Detector Configuration

Scattered light detection: Forward scattering (FSC) and backward scattering (BSC, equivalent to traditional lateral scattering SSC), both use 488/17 bandpass filters.

Fluorescence detection: equipped with 6 photomultiplier tubes (PMT), filter combination as follows:

Long pass filter: 639LP, 561LP, 487.5LP, 752LP, 685LP

Bandpass filters: FL1 450/50, FL2 525/50, FL3 600/60, FL4 665/30, FL5 720/60, FL6 785/60

This configuration supports flexible combinations of multi-color experiments.

1.3 Characteristics of Sorting System

Nozzle/chip specifications: We offer three types of microfluidic chips: 70 μ m, 100 μ m, and 130 μ m, which need to be ordered separately.

Sorting method: Supports two-way sorting (two groups of target cells+non sorted cells).

Receiving container: capable of batch sorting or single-cell sorting, supporting test tubes, 6/12/24/48/96/384 well plates, and PCR plates.

Index sorting: For each sedimentation event in the porous plate, its original parameters can be traced back on the scatter plot for subsequent single-cell analysis.

Sorting mode: Built in settings for 8 different purity/yield combinations, including a dedicated single-cell sorting mode.

1.4 Liquid Flow and Gas Path

The instrument is equipped with a power switch and pressure regulator on the back (users are not allowed to make changes on their own). The external compression pump provides a driving air pressure of approximately 6 bar. The sheath liquid tank has a capacity of approximately 2.5 liters per 8 hours of consumption rate, and the waste liquid tank needs to be emptied regularly.

Chapter 2: Physical Inspection Before Startup - Preventing Problems in Advance

Before pressing the power button, the following checks must be completed. These steps are directly related to the stability of liquid flow and the quality of sorting.

2.1 Sheath fluid tank and waste liquid tank

Lift the annular air release valve at the top of the sheath fluid tank to release the residual pressure inside the tank.

Check if the liquid level in the sheath tank is sufficient (consumption rate of approximately 2.5 liters/8 hours). If insufficient, please replace the full can: first disconnect the transparent air pipeline, then disconnect the blue sheath fluid pipeline, and remove the empty can without tilting. When placing the new can, it should also not be tilted and all pipelines should be reconnected.

Confirm that the waste liquid tank inside the liquid flow vehicle is not full. If it is close to the upper limit, please empty it.

2.2 Cleaning of Sorting Room

Open the front door of the sorting room and check for liquid splashing or salt crystallization, especially in the deflection plate area. Cleaning steps:

Wipe the inside of the transparent door, waste collector, black lateral flow sensor window, and large sensor window below the waste collector with dust-free paper soaked in ultrapure water (MilliQ).

Remove the deflection plate, clean it with ultrapure water wipes, then dry and reinstall. When cleaning the white plastic mounting block, be sure to avoid wetting the copper contacts on the back.

2.3 Liquid flow system filter and DI water tank

Open the left flow maintenance door and observe if there are any bubbles inside the sheath fluid filter (d). If there are bubbles, exhaust should be carried out after powering on.

Check if the liquid level of DI water tank (e) is too low. If insufficient, disconnect the liquid pipeline and carefully open the buckle to remove the water tank. Be careful not to let liquid drip into the air filter when unscrewing the bottle cap. After replenishing ultrapure water, put it back in place, close the buckle and reconnect the DI water pipe.

Chapter 3: Standard Startup Process - Success or Failure Determined by Order

Incorrect startup sequence may damage microfluidic chips or cause unstable liquid flow.

3.1 Hardware Power On

Firstly, turn on the power supply of the compression pump (located on the right side of the back of the equipment). This step is crucial: the compression pump must first provide about 6 bar of air pressure before proceeding with subsequent operations, especially opening the front cover of the host.

Press the power button on the right side of the front panel of the SH800 host. The green indicator light is on, and the system starts to initialize. After about 30 seconds, the screen displays "Standby".

Open the computer, log in with username fcm and password fcm.

3.2 Software Startup and Login

After SH800 completes initialization and the monitor displays "Standby", double-click the desktop SH800 icon to start the software.

Button functions in the interface:

Maintenance: Enter the maintenance program for cleaning or replacing the sample line.

Shutdown: Execute the shutdown procedure.

After entering the username and password, click Login to enter the system settings interface.

Chapter 4: Registration and Loading of Microfluidic Chips

SH800 uses disposable microfluidic chips, each chip can be used for 24 hours from registration. The registration of chips must be completed through QR code scanning.

4.1 Registration steps

Aim the QR code on the chip packaging at the built-in camera on the top of the host computer.

After the screen displays the chip information, click Next and the "Chip Loading" window will appear.

4.2 Loading Chip

Open the flip door above the front panel of the host.

If there is an old chip in the slot, it will automatically pop out and be removed.

Insert newly registered chip: Do not touch the surface of the chip. Place the side of the chip labeled with "100 μ m" (or corresponding specification) facing yourself and partially insert it into the slot. The chip loader will automatically suck the chip into place. Do not push hard to avoid damaging the chip or slot.

Click Next to enter the laser settings window.

4.3 Laser configuration and liquid flow self-test

Select the laser combination required for the experiment. The 488 nm blue laser is necessary for system setup, and other lasers can be changed later.

Confirm that the physical filter configuration is consistent with the filter mode displayed on the screen. By default, the middle option (including 405 nm laser) is selected.

After clicking Next, the instrument automatically starts the liquid flow check, which takes about 5 minutes. This process includes starting the liquid circuit, repeatedly removing bubbles from the chip, and stabilizing pressure and flow rate.

Important reminder: After the fluid flow check begins, immediately perform the defoaming operation of the sheath fluid filter (see Chapter 5). The liquid flow startup will continue in the background.

Chapter 5: Defoaming Operation of Sheath Fluid Filter

Bubbles are the main cause of unstable liquid flow, which can directly lead to sorting deviation. Even if the instrument automatically removes bubbles, it is necessary to manually check and eliminate the air inside the sheath fluid filter.

5.1 Inspection and Manual Exhaust

Open the left flow maintenance door and remove the sheath fluid filter from the fixing clip (without disconnecting the connector).

Lightly tap the side of the filter 2-3 times with your hand and observe if there are any bubbles inside. If there are no bubbles, reinstall the filter, ensuring that the arrow on the side of the filter is facing upwards, and re tighten the clip.

If there are bubbles, open the exhaust port at the top of the filter and wrap a tissue around the exhaust port to collect the spilled sheath fluid. After exhausting the air, turn the filter upside down and repeat the exhaust operation (there is also an exhaust port at the bottom).

Reinstall the filter (arrow pointing upwards) and fasten the clip.

5.2 Verification of Liquid Flow Stability

Return to the liquid flow inspection window and observe if the droplet shape is stable. If the droplet jumps, deforms, or breaks unevenly, the sheath fluid filter needs to be defoamed again until the fluid flow is in a stable straight line.

Chapter 6: Automatic Calibration - The Core to Ensure Data Accuracy

Automatic calibration uses specialized calibration microbeads (Auto Setup Beads) to optimize parameters such as laser delay and droplet deflection.

6.1 Preparation of calibration samples

Check if there are ready-made calibration tubes in the 4 ℃ refrigerator. If available and the preparation time does not exceed 7 days, it can be used directly. Remove the original calibration bead bottle, shake it thoroughly (do not vortex or shake), and add the bead solution to the scale line of the test tube.

If it has been more than 7 days, it needs to be prepared again: take a new 5 ml FACS tube, drop 10 drops of calibration beads, mark the date and liquid level on the tube wall.

6.2 Perform Calibration

Place the calibration tube into the sample holder (using the correct tube holder) and click OK.

Tip: If only analysis is performed (without sorting), click on Analyzer mode to shorten the calibration time required.

Calibration will automatically execute the four steps listed on the left side of the screen. After completing everything, check if the result shows' completed successfully '.

If successful, click OK to enter the "Create Experiment" window.

Chapter 7: Experiment Creation and Parameter Setting

After calibration is passed, a new experiment can be created.

7.1 Basic Information Input

Fill in information such as experiment name, operator, cell type, etc. in Experiment Information.

Enter the marker name and select the corresponding fluorescent dye.

7.2 Collection Parameters and Laser Selection

Select the parameter types to be collected: Area, Height, and Width, which are used for FSC, BSC (lateral scattering), and each fluorescence channel (FL1~FL6), respectively.

If not all lasers are selected at startup, you can add the required laser wavelength selection here.

After completing the setup, click on Create Experiment, and the main window will display the Experiment Explorer.

7.3 Template based Creation

Users can also create new experiments based on existing experiment templates, or directly copy recently used experiments as templates.

Chapter 8: Compensation Settings and Threshold Adjustment

Before starting to collect samples, it is usually necessary to complete fluorescence compensation and adjust detection sensitivity.

8.1 Automatic Compensation Wizard

Go to the Compensation tab and click on the Compensation Wizard to start the automatic compensation process.

Follow the guide prompts to load each single staining tube and negative tube in sequence.

After compensation is completed, the software will automatically generate a compensation matrix and apply it to the current experiment.

8.2 Detector Threshold Adjustment

Click on Detector&Threshold in the middle left corner of the screen.

Adjust the sensitivity values of each detector until a clear event cluster appears on the scatter plot.

Set appropriate thresholds (usually using FSC or fluorescence thresholds) to eliminate debris and noise.

Important reminder: Each sample tube must be filtered with a blue filter cartridge before loading. Microfluidic chips are extremely sensitive to blockage, and any particle with a diameter larger than the nozzle size will cause complete blockage.

Chapter 9: Precautions for Experimental Collection and Sorting

Confirm that the sample tube has been filtered and placed on the correct tube holder.

Before sorting, it is recommended to use non sorting mode (Analyzer mode) to observe the sample status and adjust the gate strategy.

Choose the appropriate sorting mode (purity first, yield first, or single-cell mode) during sorting.

If using a porous plate for single-cell sorting, the index sorting function can be enabled for easy traceability in the future.

Chapter 10: Shutdown and Cleaning - Irreconcilable Steps

Errors or skipping shutdown cleaning are the most common causes of permanent damage to sample lines. The following order must be strictly followed.

10.1 Start shutdown program

Select the Cytometer tab and click on [Software and Device] in the Shutdown menu.

Never click Shutdown directly in the pop-up shutdown window! Directly clicking will immediately turn off the instrument power and skip the cleaning step, which will cause the samples in the sample line to dry up and become blocked, which cannot be repaired. Users need to purchase a new sample line at their own expense.

You must click Start to start the cleaning process.

10.2 Cleaning steps

Prepare a 15 ml Falcon tube and fill it with 10 ml FACS Clean solution (bleach containing hypochlorite). Load using the corresponding sample tube holder. Attention: Do not overfill with bleach.

Software guidance completes bleach cleaning.

After cleaning, remove the bleach tube and click Next.

Prepare another 15 ml Falcon tube and fill it with 12-13 ml of ultrapure water (MilliQ). The level of DI water must be higher than the previous level of bleach to ensure that all bleach on the sample probe is thoroughly rinsed off.

Perform DI water rinse.

10.3 Turn off the instrument

After cleaning, the software asks if the host and software will automatically shut down. If using a standard sample line (not self owned), click Shutdown.

If using your own sample line and/or chip, you should cancel the shutdown and log out from the current account to return to the login interface. Then contact the facility staff to assist in retrieving the sample line and chip.

Shut down the computer.

Turn off the compression pump.

Release all pressure inside the sheath fluid tank: Lift the circular pressure relief valve at the top until no more gas is released. Long term storage under pressure may damage the instrument.

Emptying the waste liquid tank: Disconnect the pink waste liquid pipe, carefully unscrew the lid (to prevent liquid from flowing into the air filter), and pour the waste liquid into the sink. Wipe off any splashing waste liquid and disinfect with 70% ethanol. Reconnect the waste liquid tank.

Chapter 11: Common Problems and Troubleshooting

Possible causes and measures for handling the phenomenon

Droplet instability/jumping. If there are bubbles in the sheath fluid filter, perform manual defoaming in Chapter 5 and repeat if necessary

Chip blockage: If the sample is not filtered or the particle diameter is too large to clear, a new chip needs to be replaced; Ensure that all samples pass through the filter element

Automatic calibration failed due to expired calibration beads, insufficient mixing, or improper temperature. Prepare fresh calibration beads (≤ 7 days) and shake the mixture thoroughly

Sort out abnormal deflection plates that are dirty or damp. Clean the deflection plates thoroughly and dry them thoroughly; Check the voltage setting of the deflection plate

The pressure cannot be established. The sheath fluid tank is not properly sealed or the compression pump is not turned on. Check the sealing ring of the tank cover; Confirm that the power supply of the compression pump is turned on and the pressure gauge shows 6 bar

Chapter 12: Summary of Daily Maintenance Points

Before daily start-up: check the sheath fluid, waste liquid, DI water tank, and clean the sorting room.

After each sorting, strictly follow the shutdown cleaning process and do not skip it.

Weekly: Check the validity period of the calibration beads and replace them in a timely manner.

Monthly: Check the liquid flow pipeline for leaks, clean the instrument surface and fan filter.

Every six months: Deep maintenance is performed by facility professionals, including laser alignment, PMT calibration, and pipeline replacement.