Making Agar Slants (Slopes) - In Pictures

Look in here for answers to common questions and sticky threads
Post Reply
User avatar
Wolfy
Under the Table
Posts: 1565
Joined: Thu May 27, 2010 6:31 am
Location: Melbourne, Australia

Making Agar Slants (Slopes) - In Pictures

Post by Wolfy » Mon Aug 16, 2010 8:29 am

In the slant-to-starter thread someone asked for a guide to making slants/slopes, so here it is.

Agar slants or slopes are an easy way to store yeast strains.
Once inoculated with a yeast sample and refrigerated, agar slants should keep the yeast viable for months or longer.
While there is noticeable autolysis after a few months, I usually reculture slants after 1 year, others have suggested they recultured viable yeast from slants up to 4 years old.

Here is what we'll need to make our agar slants.
Clean test tubes/vials, (1.030-1.040)wort/nutrient mix, agar-agar (from an Asian supermarket), Pyrex jug and a syringe.
Image

Most slants I make are in glass 50ml test tubes, using 10ml of wort/agar in each, but for the the smaller 15ml plastic test tubes I use 5ml wort/agar.
250ml wort will make 20 larger slants (allowing for some losses).

After measuring the required amount of wort into the Pyrex jug (add some yeast nutrient if you have some), add the agar agar powder.
For the powdered agar-agar shown in the above picture, I use 1.5% agar, which works out to be about 3.75g of Agar in 250ml wort.
Image

The agar will not dissolve in the cold liquid, so you need to bring it to the boil, I do this in the microwave.
I microwave the wort/agar for about 30 secs on high, then stir, and repeat the process until it just comes to the boil.
When it gets to the boil the agar will dissolve into the wort and the mixture will be very runny:
Image

While it is still hot, measure the wort/agar mix into the test tubes, try not to drip the wort/agar mix down the sides of the tubes if you can help it.
You could tip it in from the jug but using a syringe makes it much easier:
Image
The reason I prefer agar over gelatin (which also works) is that agar exhibits hysteresis, meaning that it melts at ~85°C but it does not solidify until it cools to ~40 °C which means its easy to work with in liquid form, but once it has cooled and solidified it will not melt again (until it's heated over ~85°C).

Now we need to sterilizeour slants.
An autoclaveis the best way to do this, but not too many people have one at home, so most people sterilize our slants by moist heat in a pressure cooker or similar.
I have a (99c Ebay) pressure cooker, so I load the slants in that and 'boil' them for about 30mins at 15PSI.
Before I had the pressure cooker, I just put a couple of phone books ontop of my mini-mash-pot and did the best I could and never seemed to have any problems, but to be more pedantic I should have used the Tyndallization process:
Image

After the slants have been 'autoclaved', let them cool enough so you can handle them safely, but not enough so they start to solidify.
Take them out of the 'autoclave' and lay them on a slanted surface:
Image

Any slanted surface will do, cardboard box, tray, etc, but try to maximize the slope surface by adjusting the angle they are on:
Image

Cover with a clean tea towel (or similar) and let them cool (only takes about 1 hour now its winter), and they should solidify like so:
Image
When you fill, autoclave and lay out the slants try not to shake, tilt, or mix the slants because the wort/agar will coat the side of the tubes and make things messy, if you treat them gently you should get nice clean slopes to work with. If possible leave the caps on losely while 'autoclaving' and cooling the slants, that way there should not be too much moisture buildup inside the slants.

Since the slants should be sterile there is no need to store them in the fridge, I usually put mine into a plastic container - just to keep any contaminants off - until I need to use them. It's also not a bad idea to use tape or parafilm to seal the caps - again just to ensure they do not get contaminated.

I use the 50ml glass push-top test tubes because I have lots of them, and I got them cheap, however if I was going to buy more tubes to make slants with, I'd get screw top polypropylene 30ml vials like these ones:
Image
Last edited by Wolfy on Fri Aug 20, 2010 9:21 am, edited 1 time in total.
New stuff on my Blog: Control Box #2, Splitting liquid yeast packs, Top Cropping Yeast, Brew Day: Old 'n Brewed, Brew Day: Matrimony Mild
--------------------
I can't do everything but I'd do anything for you
I can't do anything except be in love with you
And all I do is miss you and the way we used to be
All I do is keep the beat and bad company

--------------------
Quost WebHosting - WebHosting.
Quost Domains - Cheap domain name Registration and Renewal.

User avatar
Andy
Virtually comatose but still standing
Posts: 8922
Joined: Fri Nov 18, 2005 1:00 pm
Location: Ash, Surrey
Contact:

Re: Making Agar Slants (Slopes) - In Pictures

Post by Andy » Mon Aug 16, 2010 8:44 am

Nice one Wolfy, thanks for taking the time to put that together!
Dan!

User avatar
vacant
Under the Table
Posts: 1845
Joined: Mon Aug 20, 2007 5:39 pm
Location: downstairs
Contact:

Re: Making Agar Slants (Slopes) - In Pictures

Post by vacant » Mon Aug 16, 2010 10:09 am

Nice guide.
Wolfy wrote:Since the slants should be sterile there is no need to store them in the fridge
Exactly. Leaving them out for a few days will tell you if you have one or more fails.

I was looking at jam making sites as I was thinking they do much the same thing - trying to keep content from growing unwanted stuff. After washing kilner jars in hot water and rinsing, they put them in the oven at at 140C for five minutes to sterilise. I keep wort in a 2 litre kilner jar which is also used for the final step-up so I was going to try the oven method for that and my 28ml tubes. Not sure what the melting point of the plastic screw top is, but I'll experiment on one to find out.
I brew therefore I ... I .... forget

delboy
Telling everyone Your My Best Mate
Posts: 4497
Joined: Wed Feb 28, 2007 11:33 am

Re: Making Agar Slants (Slopes) - In Pictures

Post by delboy » Mon Aug 16, 2010 11:51 am

vacant wrote:Nice guide.
Wolfy wrote:Since the slants should be sterile there is no need to store them in the fridge
Exactly. Leaving them out for a few days will tell you if you have one or more fails.

I was looking at jam making sites as I was thinking they do much the same thing - trying to keep content from growing unwanted stuff. After washing kilner jars in hot water and rinsing, they put them in the oven at at 140C for five minutes to sterilise. I keep wort in a 2 litre kilner jar which is also used for the final step-up so I was going to try the oven method for that and my 28ml tubes. Not sure what the melting point of the plastic screw top is, but I'll experiment on one to find out.
Not sure 5 mins at 140C will be up to the job when it comes to slants, due to the very high levels of sugar the jam makers have only really to worry about yeast and fungi, however when using solidified wort all sorts of nastys will want to grow in it, lots of them will have the ability to form spores which would live throught the 5 min/14Oc regimen and spring back to life when the temps had cooled.
Wet heat (ie steamed in autoclave/pressure cooker) is much more effective at killing spores, 5 mins at 121C can do the job though most leave it about 20 mins, with dry heat (oven) its recommended glassware etc be left for 2 hours at 160C.
The Daft Dog brewery.

User avatar
steve_flack
CBA Prizewinner 2010
Posts: 5606
Joined: Mon Aug 21, 2006 1:00 pm
Location: Has Left the Building.

Re: Making Agar Slants (Slopes) - In Pictures

Post by steve_flack » Mon Aug 16, 2010 11:53 am

...and shouting 'Die you b*stards!' is also ineffective I understand.
Drinking -Alt, Bitter, Slightly Smoked Mild
Waiting - More Mild
Bottled - Dubbel, Belgian Pale Ale, Baltic Porter, OFB, Golden Ale, IPA, Gold II, IPA II, Intrepid
Fermenting - OFB 3
Next Up - Stout (probably)

delboy
Telling everyone Your My Best Mate
Posts: 4497
Joined: Wed Feb 28, 2007 11:33 am

Re: Making Agar Slants (Slopes) - In Pictures

Post by delboy » Mon Aug 16, 2010 12:06 pm

Forgot to say thanks to wolfy for putting together such an excellent post im sure many will now be able to get there head around the whole slant thing now.
Liked the bit about tyndallization, im actually in the process of doing just that with some extra wort from my brewday at the weekend, its going to be getting another boil up tonight and then hopefully it should be fine at room temps until i get round to using it.
The Daft Dog brewery.

User avatar
floydmeddler
Falling off the Barstool
Posts: 3933
Joined: Mon Feb 16, 2009 10:37 pm
Location: Irish man living in Brighton

Re: Making Agar Slants (Slopes) - In Pictures

Post by floydmeddler » Mon Aug 16, 2010 1:14 pm

Good man Wolfy! I more or less follow this method but use gelatine instead. Was planning on buying some but have been very happy with the gelatine results. Will give agar a go though to experiment.

Hopefully this will inspire others to get into slanting. Have been adding droplets from pub pints to my slants to get brewery yeasts. Have also saved loads on Whitelabs strains because I only need to buy them once! :-)

User avatar
Wolfy
Under the Table
Posts: 1565
Joined: Thu May 27, 2010 6:31 am
Location: Melbourne, Australia

Re: Making Agar Slants (Slopes) - In Pictures

Post by Wolfy » Mon Aug 16, 2010 1:52 pm

@delboy, I figure if I can do it then pretty much everyone can, but hopefully lots of pictures makes it easier for others to understand how easy it is. Most of the wort I use for starters or slants etc is 'left over' wort from brew day, either the last runnings from the mash or kettle-dregs, boiled, bottled and then adjusted to the right gravity.

floydmeddler, my in-laws are visiting the UK this month, I was tempted to send some slants with them, and ask them to 'sample' various local brews so they could borrow samples of yeast for me, but instead will just have Brewlab send me a few of their strains. :)
The next sets of photos I'm working are 'streaking' yeast to isolate individual colonies (to obtain a 'pure' sample from bottle/pub obtained strains) and then freezing yeast (since once you collect a few strains reslanting gets more tedious).
New stuff on my Blog: Control Box #2, Splitting liquid yeast packs, Top Cropping Yeast, Brew Day: Old 'n Brewed, Brew Day: Matrimony Mild
--------------------
I can't do everything but I'd do anything for you
I can't do anything except be in love with you
And all I do is miss you and the way we used to be
All I do is keep the beat and bad company

--------------------
Quost WebHosting - WebHosting.
Quost Domains - Cheap domain name Registration and Renewal.

WishboneBrewery
CBA Prizewinner 2010
Posts: 7947
Joined: Sun Nov 30, 2008 9:06 pm
Location: Keighley, West Yorkshire
Contact:

Re: Making Agar Slants (Slopes) - In Pictures

Post by WishboneBrewery » Mon Aug 16, 2010 7:59 pm

Interesting stuff, how do you arrange / stand them in the Pressure Cooker?

User avatar
floydmeddler
Falling off the Barstool
Posts: 3933
Joined: Mon Feb 16, 2009 10:37 pm
Location: Irish man living in Brighton

Re: Making Agar Slants (Slopes) - In Pictures

Post by floydmeddler » Mon Aug 16, 2010 10:50 pm

Wolfy wrote: The next sets of photos I'm working are 'streaking' yeast to isolate individual colonies (to obtain a 'pure' sample from bottle/pub obtained strains)...
I've used a petri dish and streaked for the first time a few days back. Streaked the slurry from a bottle of Chimay. sadly nothing has appeared yet. Will try again another time.

Streaked some Duvel slurry onto a slant and it's showing signs of life. :-)

Yep, photos of this process would be great. Also, I'm happy to post you slants if there are yeasts you want super bad. Sure we could work something out.

User avatar
Gricey
Under the Table
Posts: 1084
Joined: Sat Jul 03, 2010 9:07 pm
Location: Sheffield, UK
Contact:

Re: Making Agar Slants (Slopes) - In Pictures

Post by Gricey » Mon Aug 16, 2010 11:38 pm

This is amazing. Post more! :mrgreen:
Bad Panda Brewery
Fermenting: FV1: AG#18 English IPA FV2: AG#19 Summer Dunkelweizen
Conditioning: AG#16 Chimay Reddish, AG#17 Amarillo Brillo
Maturing: AG#05 B.O.R.I.S.: Bricksh*tter Oatmeal Russian Imperial Stout - ready 01/10/11, AG#07 Monkey Shot! IAPA - ready 16/06/11 maybe
Drinking: AG#11, AG#14, AG#15
Planning: AG#20 Summer Hefeweisen, AG#21 Saison Brettre, AG#22 Simcoe Poisoning Red IPA, AG#23 Oatmeal Stout

User avatar
Bobba
Lost in an Alcoholic Haze
Posts: 563
Joined: Thu Mar 19, 2009 12:24 am
Location: Cambridge

Re: Making Agar Slants (Slopes) - In Pictures

Post by Bobba » Mon Aug 16, 2010 11:58 pm

=D> Thanks for taking the time to put this up wolfy. This is definitely something I'm thinking of. How do you find the yeast from slant compares to reculturing from a bottle?

FV: -
Conditioning: AG34 Randy's Three Nipple Tripel 9.2%, AG39 APA for a mate's wedding
On bottle: AG32 Homegrown Northdown ESB, AG33 Homegrown Cascade Best
On tap: -
Garden: 2x cascade, 2x Farnham whitebine (mathon), 2x northdown, 1x first gold

User avatar
Wolfy
Under the Table
Posts: 1565
Joined: Thu May 27, 2010 6:31 am
Location: Melbourne, Australia

Re: Making Agar Slants (Slopes) - In Pictures

Post by Wolfy » Tue Aug 17, 2010 6:31 am

pdtnc wrote:Interesting stuff, how do you arrange / stand them in the Pressure Cooker?
As per the pictures. ;)
The flat bottom tubes/vials I just stand in the cooker, on a plate, that is raised off the bottom, by resting it on some ceramic chop-stick-holders, then fill with water to the level of the plate.
The curved/pointed tubes are more difficult, so I just put them in the Pyrex jug and then put that in the cooker, again raised off the bottom a little.
floydmeddler wrote:Yep, photos of this process would be great. Also, I'm happy to post you slants if there are yeasts you want super bad. Sure we could work something out.
I've been trying (to work out a method of) streaking onto the slants, the media is the same, but there is just less surface than petri dishes. However, test tubes/vials are also cheaper and easier to obtain for most people.
Pending no issues with Australian Customs or Quarantine, Brewlab will be sending some of their yeast over here, so that should cover many of the yeasts you can find in pubs/bottles there I think - but thankyou for the offer. :)
Bobba wrote:How do you find the yeast from slant compares to reculturing from a bottle?
Since I use the entire slant each time I make a starter (as per the other thread), I use much the same procedure as if i was reculturing from a bottle.
With a slant I tip about 5ml of fresh wort into the slant and scrape off the yeast before stepping that up to 20ml, from a bottle - depending on how much sediment there is - I'll usually start with the 20ml wort and build the starter from there.
If you can reculture from a bottle, slants are easier IMHO - at least you know the yest should be viable and in fairly good condition.
New stuff on my Blog: Control Box #2, Splitting liquid yeast packs, Top Cropping Yeast, Brew Day: Old 'n Brewed, Brew Day: Matrimony Mild
--------------------
I can't do everything but I'd do anything for you
I can't do anything except be in love with you
And all I do is miss you and the way we used to be
All I do is keep the beat and bad company

--------------------
Quost WebHosting - WebHosting.
Quost Domains - Cheap domain name Registration and Renewal.

User avatar
johnmac
Under the Table
Posts: 1384
Joined: Fri Jan 27, 2006 6:00 pm
Location: Shropshire

Re: Making Agar Slants (Slopes) - In Pictures

Post by johnmac » Wed Aug 18, 2010 9:36 am

Is there any harm in saving your old used slants and melting them down to use again?

User avatar
floydmeddler
Falling off the Barstool
Posts: 3933
Joined: Mon Feb 16, 2009 10:37 pm
Location: Irish man living in Brighton

Re: Making Agar Slants (Slopes) - In Pictures

Post by floydmeddler » Wed Aug 18, 2010 12:26 pm

johnmac wrote:Is there any harm in saving your old used slants and melting them down to use again?
Interesting question.

Post Reply

Return to “How-To Guides, FAQs and Stickies”

Who is online

Users browsing this forum: No registered users and 1 guest