This BYOC was designed to achieve one of my goals I had set upon joining the ISAC SRL EL program. The purpose of the BYOC is to act as an additional aid to spark interest when teaching cytometry to students. It has been designed to be affordable ($500-$1500), 3D printable and fun to build. I have included the build list, instructions, notes and highlight there are many opportunities for improvement/modification (I can attest to this with my experience over the last 6 years). 

The BYOC Workshop was first held @ ACS 2016 in Sydney, Australia, @ ACS 2018 in Adelaide, Australia and conjointly with the Garvan Flow Facility @ ACS 2019, Melbourne, Australia. This page was updated with the plan to host the workshop @ ACS2022 in Melbourne. 

Overview

The cytometer uses a <$50 adjustable 5-25mW laser (interchangeable) that is directed onto a jet in air stream. The stream is ejected throughout a 100um nozzle that is assembled using luer locks, cogs and HPLC tubing to allow injection of sample in a laminar flow. The laser is focused onto the stream and signals are detected using a 25mm, 25mm focal lens that directs the emitted/scattered light onto the FSC/SSC/FL detectors. A diagram of the light path from excitation is shown in the diagram below. The signals are detected with a APD/Photodiode or PMT and then converted to a voltage output using a pre-amp. The signals can be viewed using an oscilloscope or can be displayed on a bivariate dot plot using a ADC connected to a computer. 

3D Printable Parts (2022 updates)

Steering laser mirror mount - https://www.thingiverse.com/thing:3523131

Flow cell - https://www.thingiverse.com/thing:3902321

Detector - https://www.thingiverse.com/thing:3902266

Mounting - https://www.thingiverse.com/thing:3902335

Sample tube mount - https://www.thingiverse.com/thing:3902328

Flow control - https://www.thingiverse.com/thing:3902432

Laser mount - https://www.thingiverse.com/thing:3902366

Notes

• The laser interception point as of 2022 has been fixed at 127mm from the base. This 'fixing' allows for a simpler setup and easier modifications. 
• Using the relatively affordable 'blackridge' air regulator from Supercheap auto, with a 0.1um PES filter in line, dial 3 full rotations and then dial back a 1/2 rotation of the air pressure regulator valve for this project. This provides approximately 2-3psi that is hard to observe on the blackridge air regulator. 
• When focusing the laser it is important to focus it so that there are no striations seen when centered. Striations indicate a focal point either behind or infront of the stream. An optimal focus will have striations briefly when aligning the laser stream and it is crossing the edges but will result in no striations when centered. 
• The microscope used in this project is a modified kids toy without any optical filters. To view the core stream interception point ensure the microscope is angled so that stray laser speckle does not overwhelm the scope.  
• The testing rig LED can be run from an external pulse generator. >1000 pulses per second work well for the preamplifier circuit design. Try increasing the speed of the flash if there is a negative pulse. 
• 9x9V for Photodiodes
• 86V saturates APD, 83V just detects, 85 less.

Notes - other references

mmHg to PSI

**Divide by 51.715 to obtain PSI**

Bill of materials - 3D Printed Version (acrylic assists can be interchanged)

Item	Qty	Purpose	Sources	Cost ($AUD)	Notes	Image
600x600 tile	1	base	Local tile supplier	Free (tell them what you are making)	Aim for a free marble tile
600x600 steel sheet	1	build platform	Bunnings (Sydney)	$26	Ensure it is magnetic. Thickness does not matter, thinner normally means cheaper. I recommend sign steel. Have some fun and wobble it ;p
0.3M7TIA gear	2	holds HPLC in the centre of luer lock	Aliexpress	<$5 for 20	https://www.aliexpress.com/item/33000449925.html?spm=a2g0s.9042311.0.0.86a84c4dqSRqUr
450 extension springs	19 (2 detectors), additional 6 per detector	provides pulling force to mirror, laser, lenses	Bolts & Industrial (Sydney) Century Springs (US)	$4	We need to buy it from a reseller. Spring Catalogue: https://www.centurysprings.com.au/products/prod_catalogue.htm Can also buy about 10x cheaper from ebay.
M3 socket screw	bag	used as screws to put the whole thing together	RS components or ebay
M3 nut	bag	Ensure outside dimensions are 5.5mmx5.5mm	RS components	$13.80	RS PRO, M3 5.5mm Steel Square Nuts, Bright Zinc Plated Finish  837-262
25mmx25mm optics	2-3	focus light onto detector from intercept point	Edmund optic	$52.80	Stock #32-490 https://www.edmundoptics.com.au/p/25mm-dia-x-25mm-fl-mgfsub2sub-coated-double-convex-lens/2375/
5mmx25mm optics	1	focus laser light onto stream	Edmund optic	$61.50	Stock No. #87-913 https://www.edmundoptics.com.au/p/50mm-dia-x-250mm-fl-mgfsub2sub-coated-plano-convex-lens/30185/
100um nozzle	1	eject stream	Intertronics or FISNAR	38 pounds	https://www.intertronics.co.uk/product/preci-tip-precision-dispensing-tips/ IDMSPREC-M100-NS
Luer lock connectors and 1/4inch BSP to barb fittings	1	to connect to nozzle	Bunnings & Kinesis Australia & USA KENT		I will need to seperate this column a bit more - information will be provided during the workshop.
30mm x 10mmx 3mm	20	mount items	Ebay	$18	20pcs N50 Strong Block Cuboid Rare Earth Neodymium Magnets 30mm x 10mm x 3mm https://www.ebay.com.au/itm/20pcs-N50-Strong-Block-Cuboid-Rare-Earth-Neodymium-Magnets-30mm-x-10mm-x-3mm/171206332711?hash=item27dcb15127:m:mkKfgJMNa9H2_6E0RsAkhxQ&frcectupt=true
Circular magnet		To mount APD instead of photodiode
20L Drum	2	used to create a vacuum	recycle bin	free	to make see instructions
Regulator	2	adjust pressures	supercheap auto	$30 each  or $30 for 2	https://www.supercheapauto.com.au/p/blackridge-air-regulator/340078.html or  https://www.ebay.com.au/itm/1-4-Inch180PSI-Air-Compressor-Pressure-Regulator-Gauge-Switch-Control-Valve-Kits/264442958026?ssPageName=STRK%3AMEBIDX%3AIT&_trksid=p2057872.m2749.l2649
APD or Photodiode	1 minumum	convert photons to current	many sources including old instruments	$15 each	APDs used are AD500-8 TO52 package or BNC photodiode package
Laser 532nm (or equivalent)	1		AIXIZ lasers	$38.50USD	https://www.aixiz.store/product-page/532nm-5-25mw-adjustable-power-3vdc-line-cross-attachable-ad-532-25adj
Electronics shopping list (per detector)
Prototyping board	1			$8.19	https://core-electronics.com.au/solder-able-breadboard.html
RJ45 breakout	2			$1.33	https://core-electronics.com.au/breakout-board-for-rj45.html
LT1364CN8#PBF-ND	2	opamp used for I>V and for amplification	DIGIKEY	#13.73 each	https://www.digikey.com/product-detail/en/linear-technology-analog-devices/LT1364CN8-PBF/LT1364CN8-PBF-ND/891887
Cat7 RJ45 cable	1			$8	Amazon/Ebay
Adjustable 10^5 25 turn resistor				<$1	Jaycar (generic electronics parts store)
Adjustable 10^3 25 turn resistor				<$1	Jaycar (generic electronics parts store)
1N4148 diodes	4			<$1	Jaycar (generic electronics parts store)
10pF ceramic capacitors	2			<$1	Jaycar (generic electronics parts store)
10K resistor (1% tolerance)	3			<$1	Jaycar (generic electronics parts store)
100K resistor	2			<$1	Jaycar (generic electronics parts store)
1K resistor	1			<$1	Jaycar (generic electronics parts store)
22K resistor	1			<$1	Jaycar (generic electronics parts store)
0.22uF ceramic cap	1			<$1	Jaycar (generic electronics parts store)
1uF ceramic cap	1			<$1	Jaycar (generic electronics parts store)
2 pin molex pcb header 2.54mm pitch	1			$0.20	https://www.jaycar.com.au/2-pin-0-1-straight-locking-header-2-54-pitch-single/p/HM3412
3pin molex header female 2.54mm pitch	1	used to connect to APD		$0.55	https://www.jaycar.com.au/3-pin-0-1-header-with-crimp-pins-2-54-pitch/p/HM3403
hookup wire and crimpers				$0	Spares in every household
9V battery connectors	4	buy different types of connectors to ensure flat 80-90V power supply		$1.40
8 pin IC housing	2	mount for  IC chips		$1.25	https://www.jaycar.com.au/8-pin-gold-insert-machined-pin-ic-socket/p/PI6452
4 or 6 way DIP switch	1	used to turn power on / off		$1.65	https://www.altronics.com.au/p/s3050-4-way-dip-switch/
shielded RJ45 port	2	needed to suppress noise		$1.75	https://www.altronics.com.au/p/p1538-oupiin-8p8c-rj45-modular-socket-shielded-pcb-mount/
Differential pressure regulation
pressure transducer - 24PCAFA6D	1	accurate pressure differential between sample/flow cell	Element14	$61.89	https://au.element14.com/honeywell/24pcafa6d/pressure-sensor-0-1psid/dp/731638?st=24pc%20pressure%20sensor 0-1psi - 0-45mV - 2002508.pdf
OPAMP - breakout for pressure transducer	1	simple 2 stage amplification of 100	Core electronics	$9.90	BOB-09816  100x gain  https://core-electronics.com.au/opamp-breakout.html
LCD keypad	1	pressure output monitor	Core electronics	$16.50	https://core-electronics.com.au/lcd-keypad-shield-for-arduino.html DFR0009
arduino and case	1	arduino uno	Core electronics	$39.95 + $4.70	CE05634 A000066
Aluminum laser mount and heat sink	1	holds and cools the laser	Aixiz laser supplies	USD$3.95	12X30mm aluminum mount for AixiZ 12X30mm units [AIX-ALM-MT-1]
6mm nuts	many	mounts with 3mm threads	RS components	$12.23 pkt of 100	RS PRO, M3 5.5mm Steel Square Nuts, Bright Zinc Plated Finish 837-262
Molex 2 way connector and crimp	1x for laser power source	Allowing the disconnection renders the laser and power supply modular.	RS components
Cable ties	many	Need I say more			100mm x 2.5mm
Polulu fine control stepup/down  voltage regluator	1 per APD detector	9V batteries are coarse adjustment - this gets you finer control	core electronics	$27.15	https://www.pololu.com/product/2869

Instructions 

Setting up the laser

Most standard cytometers use a laser as a light source. The BYO cytometer is no different. Read on to mount your laser!

• Insert the laser of choice into the aluminum heat sink and hand tighten the screw to lock the laser in.
  
• Here are 2 compatible lasers (left is a 405nm, right is a 532nm laser available from aixiz.com.
  
• Insert 2x5.5mm square nuts into the laser mount nut sockets and attach the laser mount using 2xM3 10mm socket screws
  
• Insert 2 rectangular magnets into each base to secure the laser to the steel base.
  
• Attach a 2.5mm ~100mm cable tie to secure the laser power cables. This is to prevent them falling off with too much movement. 
  
• Cut off the excess on all cable ties going forward
  
• Crimp on a 2 way molex connector using a crimper
  
• Attach the connector and use a scapel to expose the laser control potentiometer (the yellow heat shrink is what you need to cut back). 
  
• The laser is now ready!

Making a flow cell...

• obtain male to male luer lock (transparent is great), and 2 small 0.3M7TIA gears
  
• Setup the luer lock to hold the gears - this will be used to hold the PEEK tubing. 
  
• Insert the precut peek tubing 
  
• Connect the remaining luer lock connectors followed by a luer lock to barb connector as shown
  
• Using 2 Y connectors, tubing and luer lock to bar connectors create the following Y connector that will join the stiff main flow cell. 
  
• Pass the peek tubing up through the female luer lock connector.
• Create a sheath bleed point and transition to silicon tubing as shown.

Let's vacuum!

The instrument uses a vacuum to draw away excess liquid. There are a few variants to the vacuum (including a gravity version that I absolutely love, and not only because I designed it, and it lasts ~10 minutes).

This version is the electric vacuum pump version. 

• Obtain a vacuum pump (listed above). 
• placeholder
• Obtain a wine bottle with a aluminum lid, preferably a top shelf one . Utilise wine thoughtfully. 
• To drill the holes we start with a guide drill bit ~1-2mm, drill our holes and then follow through with a hole size that is about ~2mm smaller than the tubing. I've found a 4mm drill bit works well for the ~6mm Tygon tubing obtained for this project. I'll post exact specifications above. The wine bottle lid requires 2 holes, 1 to draw out the vacuum and 1 to lead the liquid waste in.