Prevent autoclave water from seeping into pipette tip boxes

When I want to autoclave pipette tip boxes, I would place these boxes at higher level during autoclave. Placing these boxes (or other materials) near to the perforated platform just above the autoclave water would invite the "dirty" autoclave water (sometimes we tend to change the water once every two days or more) from seeping into the tip boxes. The brownish (probably due to spilt media) water will seep into the boxes and when dried, the tips will have a yellowish to brown stain covering them later.

When the tip boxes are placed neat to the perforate platform just above the autoclave water, then during the autoclave, the dirty water will thrash inside the autoclave and some of the water will seep into the tip boxes via cracks. That will contaminate these boxes.

The dirty water might contain impurities that will inhibit sensitive reactions or worst, contaminate stock solutions, enzymes, and expensive reagents.

Not all labs would use commercially packed filtered tips and we do use sterilized tips for dispensing expensive reagents. It is very important to properly autoclave tip boxes so that no contamination from the autoclave water will occur.

I had my fair share of dirty tips due to the above type of autoclave machine.

Just to share.

DIY Brine shrimp (or Artemia) hatchery

Introduction
Brine shrimp (or Artemia salina) is a crustacean used as a food source for fry and fish. In a zebrafish facility, brine shrimp is used as a intermittent feed for the fry and adults. Shrimp hatchlings at less than two days old contain yolky reserve rich in nutrients for fry and adult fish (feeding them artemia increases better egg production in females).

Materials

• Brine shrimp eggs. Don't get those decapsulated eggs that will not hatch BUT used directly to feed fish instead. For bulk order, you can visit BrineShrimpDirect (I have not dealt with them so any question do contact them directly). In eBay, the cost is  about SGD 2.20 per 10 g of eggs. Normally, I will use 2 tbsp per 1 litre sea salt water and allow it to hatch overnight with aeration.

Brine shrimp eggs from eBay seller fishmanofrobertsbridge.

• Aquarium sea salt mixture (weighing 1.36 kg) available at eBay from timholliday at approx. SGD 36.00 (inclusive of shipment). About 20 to 30 g salt per liter of water is required to hatch brine shrimp (but best would be to follow the instruction from shrimp egg producer).

Sea salt from eBay seller timholliday.

• Shrimp hatchery set. You can DIY by using a Coca Cola 2 liter bottle, aquarium tubing, and plastic hollow balloon sticks (see fig below).

 

DIY brine shrimp hatchery. Instead of having a nozzle at the bottom to drain out the hatched shrimp, I prefer using the balloon stick connected to tubing to siphon off the shrimps from the vessel. That way, I will not be collecting unhatched eggs settled to the bottom as I would by using the bottom outlet (in commercial setting). 

Once the unhatched eggs are settled to the bottom and the empty cyst floating at the surface, I would usually use the hollow stick/straw connected to tubing to siphon out the shrimp (indicated red). A muslin cloth is normally used to net the shrimp and added back to fish water (at a reduced salt concentration) before feeding.

Different stages of maturity of the brine shrimps. New hatchlings are preferred due to their high yolk content, whereas older shrimps might have loss some yolky storage. The unhatched eggs are shown here.

 For commercial hatchery, you can search for them from eBay. Just key in brine shrimp hatchery and you will get returns.

Notes

• Hatching the eggs is good for maximum three days. After that, the shrimp will lose its yolk reserve and would also have lost their nutritious value. If you need to keep shrimp for more than three days, it is best to feed the brine shrimp in order to keep them alive.
• Don't feed too much shrimp to your fish because this food is considered fatty. Feed the fish once per day and add in dry food for the other two feeding per day (total 3 feeding).
• Fish will always be hungry. So, don't allow demand feeding because there will always be demand. Fish will die of obesity-related complications!

DIY material

Hollow balloon sticks (get white ones if preferred). Fig from www.balloonsandweights.com.

Muslin cloth or cloth coffee filter (you can get this from Aliexpress or shops)

Protein concentration, membrane based, Molecular Weight Cutoff aka Nominal Molecular Weight Limit

Protein concentrator based on membrane and centrifugation works by removing biomolecules at determined molecular size/weight. For example, a membrane-based protein concentrator such as Centriprep Centrifugal Filter Unit with Ultracel-50 membrane (Merck Millipore cat# 4311) has a protein cutoff point at 50 kDa. The term used can be  Molecular Weight Cutoff (MWCO; refer to Wikipedia) or Nominal Molecular Weight Limit (NMWL) but they mean similar thing, that proteins at or above 50 kDa is trapped/concentrated whereas molecules at <50 be="" br="" kda="" removed.="" will="">

Items for zebrafish injection

Capillary tube ( www.wpi-europe.com/products/glass/patch-clamp-glass-capillaries.aspx).

Pipette man has carpal turner syndrome...

Have you ever experience a pipette man turn creaky on you? I had. The "piston" just wouldn't slide smoothly and sometimes got stuck. This is really irritating especially when you required delicate pipetting to prevent removing pellet or damage zebrafish embryos.

A lubricant is needed, whereas I, a mocha.