Laboratory Techniques

This page will focus on home lab and small brewery lab techniques.

Equipment

General (links to other pages)

• Microscope
• pH Meter

Density Meter

Anton Paar Portable Density Meter

UV Plate Cooling Cabinet

Using a UV sterilization cabinet can not only help cool/sterilize freshly poured plates but it can be used to UV sterilize other things as well. The build itself is quite simple. You will have to do some basic wiring.

Parts List - Light Ballast https://www.amazon.com/gp/product/B00AB32J7S/ref=oh_aui_detailpage_o04_s01?ie=UTF8&psc=1 Lamp Mount https://www.amazon.com/gp/product/B0036ZA966/ref=oh_aui_detailpage_o04_s01?ie=UTF8&psc=1 4 pin connector https://www.amazon.com/gp/product/B003B92NP2/ref=oh_aui_detailpage_o04_s01?ie=UTF8&psc=1 UV Light https://www.amazon.com/gp/product/B001HB3E2W/ref=oh_aui_detailpage_o06_s00?ie=UTF8&psc=1 Plexiglass https://www.amazon.com/gp/product/B019D0DUDQ/ref=oh_aui_detailpage_o03_s00?ie=UTF8&psc=1

Orbital Shaker

An orbital shaker is a laboratory device used for mixing substances or maintaining movement of fluids. Maintaining movement of liquids has been shown to help some microorganisms grow. For example, running a shaker at 80 RPM for Brettanomyces starters is an effective way to grow this genra (see Brettanomyces starters) . For a home orbital shaker example, see Example of a Home Lab Orbital Shaker.

Growth Media

Lactobacillus/Pediococcus

See Rogosa SL Agar

MRS Media

Chemical	Usage Amount
Dextrose	20 grams
Peptone	10 grams
Beef Extract	8 grams
Yeast Extract	4 grams
Sodium Acetate	5 grams
Dipotassium Hydrogen Phosphate	2 grams
Ammonium Citrate	2 grams
Manganous Sulfate Tetrahydrate	0.05 grams
Magnesium Sulfate Heptahydrate	0.2 grams
Distilled/De-ionized Water	Fill to 1000 ML

Saccharomyces

A wide variety of media can be used for Saccharomyces. Bromocresol Green can also be added to these media, as in the commercial WLN formulation. Most Saccharomyces cannot metabolize this dye, causing the colonies to stain green. Chloramphenicol can also be added to eliminate bacterial growth. Although not all of these media are specifically for your average brewers Saccharomyces, most strains should have no issues growing.

YPD Media

Chemical	Usage Amount
Yeast Extract	10 grams
Peptone	20 grams
Dextrose	20 grams
Agar(optional)	15 grams
Distilled Water	Fill to 1000 ML

MYPG Media

Chemical	Usage Amount
Malt Extract	3 grams
Yeast Extract	3 grams
Peptone	3 grams
Dextrose	10 grams
Agar	15 grams
Distilled Water	Fill to 1000 ML

Sabouraud Media

Chemical	Usage Amount
Cycloheximide (Optional)	10 mg
Chloramphenicol (Optional)	0.5 grams
Peptone	5 grams
Dextrose	20 grams
Agar	15 grams
Distilled Water	Fill to 1000 ML

Freezing Media

Chemical	Usage Amount
Glycerin	50 grams
Ascorbic Acid	15 grams
Liquid YPD/MYPG	Fill to 100 ML

Beef Broth Media

Chemical	Usage Amount
Beef Broth(No preservatives)	500 mL
NaCI (can substitute non-iodized or sea salt)	50-200 grams
Peptone	5 grams
Dextrose	10 grams
Agar(optional)	17 grams
Distilled Water	Fill to 1000 ML

Wild Yeast Screening Media

Chemical	Usage Amount
Peptone	5 grams
Yeast Extract	3 grams
Malt Extract	3 grams
Dextrose	5 grams
CuSO4	310 mg
Distilled Water	Fill to 1000 ML

Brettanomyces

A few different medias can be used to isolate Brettanomyces but DBDM media is commonly used. WLD with additions of cycloheximide can also be used.

DBDM Media Recipe

Chemical	Usage Amount
Yeast nitrogen base (YNB)	6.5 grams
Ethanol	4% v/v
Cycloheximide	10 mg
p-coumaric acid	100 mg
Bromocresol Green	22 mg
Agar	20 grams
Distilled Water	Fill to 1000 ML

WLD Media Recipe

Chemical	Usage Amount
Cycloheximide	4 grams
Yeast Extract	4 grams
Pancreatic Digest of Casein(Pepton)	5 grams
Dextrose	50 grams
Monopotassium Phosphate	.55 grams
Potassium Chloride	425 mg
Calcium Chloride	125 mg
Magnesium Sulfate	125 mg
Ferric Chloride	2.5 mg
Manganese Sulfate	2.5 mg
Bromocresol Green	22 mg
Agar	20 grams
Distilled Water	Fill to 1000 ML

Misc/Other

• MTF tips on autoclaving wort extract based media in such a way that prevents precipitation and darkening.
• For techniques on separating yeast strains from themselves and from bacteria, see Yeast/Bacteria Isolation.

Reference on DBDM: https://www.facebook.com/groups/MilkTheFunk/permalink/1805210829507123/?comment_id=1805397166155156&comment_tracking=%7B%22tn%22%3A%22R%22%7D

Storage

• For lactic acid bacteria storage, see Lactobacillus Storage.
• For Brettanomyces storage, see Storing Brettanomyces.
• For S. cerevisiae storage, see Saccharomyces Storage.

Propagators

• MTF thread on DIY and carboy-sizzed propogators for small breweries.

Justin Amaral's 4.5 BBL Propagator

The propagator below has a 14 gallon capacity, allowing a prop up of 4.5 BBL. This same set up can be used on any fermenter but is bested used on ones that can hold 5 PSI and up. The seal on the below fermenter can hold around 5 PSI but clamps are also used on the lid to ensure it can hold up to 10 PSI. It essentially acts as a large stir plate while allowing to trickle in either O2, NO2 or CO2 depending on what your propagating. Ideally you'd also use a non magnetic drive pump but its not essential.

Below is the draw up of the build out for this prop up tank. As you can see it uses quick disconnects but you can use tri-clover connects as well. It also uses a diffusion stone that connects into one of the ferrules allowing you to trickle in gases. The picture shows a diffusion stone directly into the fermenter but its since been adapted as shown in the other pictures below to allow the diffusion stone to be behind a valve so its only exposed when needed. It also uses a whirlpool connector that goes into the fermenter creating the whirlpool when the pump is on:

Below you can see the 4 way SS T valve at the top of the fermenter allowing for a blow off tube, sterile transferring of yeast(the yeast has to be in a container with a tri-clover valve for this), and the hose coming from the kettle:

Below is the valve set up. The top valve has the whirlpool connect, the middle has the diffusion stone behind the valve, and the bottom has an elbow with a valve:

Techniques

(Videos provided by Bryan of Sui Generis blog and Zach Taggart of 42 North Brewing Company.)

Making Agar Plates

• Making WLD plates:

Yeast/Bacteria Isolation

• Mark Trent details his method for isolating Brettanomyces from Saccharomyces from dregs or other mixed cultures.
• See Wild Yeast Isolation.
• See isolating Lactobacillus.

Making Your Own Media

Although media can be bought pre-made, you can also make these media yourself. Media can be either sterilized via an autoclave/pressure cooker or using sterile filtering. Keep in mind it can be difficult however to sterile filter some items such as yeast extract, peptone and brewer's grade DME. Because of this some just autoclave parts of the media while sterile filtering the rest into the sterilized media.

It is crucial you use aseptic technique with your media once it is sterilized to prevent any contamination. If you are storing extra media for later use make sure to remake it every month or 2 if unused.

Shipping Cultures

Example of DeWayne Schaaf's packaging for shipping cultures.

DeWayne Schaaf recommends the following procedures for shipping cultures with the USA states and territories :

1. Use 15ml centrifuge tubes from Cynmar LLC - Wine & Brew. They are great quality and have a very low rate of leakage, especially when combined with electrical tape.

1. Place each liquid filled tube into its own snack sized baggie to minimize cross contamination if they do happen to leak.

1. Each of these smaller bags will be placed into a quart Ziplock.

1. My main shipping container are Uline Poly bubble mailers. I don't tend to use ice packs as I've found them unnecessary when shipping in favorable temps (use ice packs during the summer).

1. USPS is my preferred shipping method. Using first class shipping, my packages typically take no more than 3 days to reach somewhere in the USA and cost right around $3, and that includes a tracking number. Shipping rates are roughly the same for any USA state or territory. The 48 continental states, Alaska, Hawaii, Puerto Rico, and Guam all cost roughly the same.

For 5 gallon pitches or cultures that may still be a bit active creating cO2 Soda Preform tubes work very well. These are the same type of tubes White Labs uses for their homebrew pitches. https://www.amazon.com/Soda-Bottle-Preforms-Caps-30/dp/B008MB1QNY/ref=sr_1_1?ie=UTF8&qid=1504524577&sr=8-1&keywords=soda+test+tubes

See also:

• Bryan of Sui Generis Blog guide to shipping yeast (recommended for larger quantities).

Historical

• "Pasteur and the beer of national revenge", by Lars Garshol (how Louis Pasteur proved that microbes were alive in 1857, and other discoveries).
• "Emil Chr. Hansen and the yeast revolution", by Lars Garshol (how yeast was first isolated from other strains in 1883).

See Also

Additional Articles on MTF Wiki

• Microscope
• PH Meter
• Wild Yeast Isolation
• Titratable Acidity
• Saccharomyces
• Brettanomyces
• Pediococcus
• Lactobacillus

External Resources

• BasicBrewing interview with Zack Taggart, lab manager at 42 North Brewing, on how to set up our own homebrew lab to analyze yeast health and count cell population (part 1) and isolating, propagating, and storing yeast and bacteria (part 2).
• "Brewers' Laboratory Handbook: Brewing Without the Blindfold™", by Brewing Science Institute.