Culture Journal, Species: Isochrysis galbana

Author Message
JimWelsh
  • Total Posts : 1426
  • Reward points : 1486
  • Joined: 1/22/2010
  • Location: Angwin, CA, US
Culture Journal, Species: Isochrysis galbana - Thursday, September 30, 2010 2:27 AM
Culturing Journal DataSheet
This first post should be updated regularly to include new information as events take place or changes are made to your system




General
Species:  Isochrysis galbana
Species description:  Motile phytoflagellate 4-6 microns
Culture source (link if possible):  SeahorseSource.com
If algae, CCMP # (ref http://ccmp.bigelow.edu/ ):  CCMP 462, 1323, 1611

Culturing Vessel Details
Salinity:  1.019
Temperature:  Ambient room temp; 72 - 74 F NOTE:  The culture itself actually gets up to abou 78 - 80 F.
Vessel description:  2 liter borosilicate media culture jars
Lighting description:  1 warm white 40W fluorescent + 1 daylight 40W fluorescent.  NOTE:  That was the old phyto station.  The new one is using 2 x 6500K 21 Watt T5 bulbs in each fixture (one fixture per shelf).
Lighting cycle:  18h on / 6h off
Aeration description:  vigorous

Methodologies
Split methodology:   Split 4:1 (500 ml of dense culture + 1500 ml of fresh sterilized water)

Culture medium description:  1200 ml of recycled tank water + 300 ml of fresh RO water microwaved to 85 C (185 F), plus 0.75 ml of FAF Micro Algae Grow (sufficient to make a Guillard F/2 solution for 2000 ml total volume).

Cell count:   At its densest, the cell count is roughly 15,000,000 - 20,000,000 cells/ml, based on Secchi readings and charts.

Reference links:  

Additional Information
Notes: 



You will be required to provide photographic evidence and as much detail as possible about your project in this thread.
If your thread does not contain detailed enough photos  and information the MBI Council will not be able to approve your reports.

<message edited by JimWelsh on Tuesday, March 27, 2012 10:05 AM>

JimWelsh
  • Total Posts : 1426
  • Reward points : 1486
  • Joined: 1/22/2010
  • Location: Angwin, CA, US
Re:Culture Journal, Species: Isochrysis galbana - Sunday, August 14, 2011 8:42 PM
I thought I'd dust this journal off and update it.  My current Isochrysis culture was established on 20 October 2010, with a starter of C-Iso from SeahorseSource.com.  It is the light yellow culture on the right-hand side of the middle shelf in the attached photo.  The dark brown cultures in this photo are my older T-Iso cultures, which I no longer have.  The C-Iso culture has been in continuous culture in my phyto station ever since, and is still going today.
 

<message edited by JimWelsh on Sunday, August 14, 2011 9:02 PM>
Attached Image(s)

chuenwe
  • Total Posts : 97
  • Reward points : 113
  • Joined: 7/9/2011
  • Location: Huntington Sta, NY, US
Re:Culture Journal, Species: Isochrysis galbana - Sunday, August 14, 2011 8:44 PM
Sweet!  I gotta buy one from SeahourseSource to try again.  I failed miserably using the dish from florida farm.
What pump is that?
 

JimWelsh
  • Total Posts : 1426
  • Reward points : 1486
  • Joined: 1/22/2010
  • Location: Angwin, CA, US
Re:Culture Journal, Species: Isochrysis galbana - Sunday, August 14, 2011 9:02 PM
Here is a picture of the C-Iso culture today.  I just split the dark culture into two new cultures this morning.  Following my standard phyto protocol, I took 1200 ml of used tank water, plus 300 ml of fresh RO water, in each media jar, plugged the tops with a foam stopper, and microwaved two such media jars in an 1100 watt microwave for 25:80  (Just pushed the keys from the top down the middle column on the keypad, = 26:20).  This amount of time brings the temp up to about 200 F.  It really just needs to get over 185 F, but I like to be certain I've killed everything.  I let it cool to room temperature, and then add roughly 0.75 ml of FAF Micro Algae Grow, which approximates F/2 strength medium.  I then remove the stopper from a fresh media jar, remove the stopper and airline from an existing culture that is dark chocolate brown, and quickly pour about 500 ml from the dark culture into the fresh media.  I then take a prepared clean stopper and airline (either entirely new, or else a used airline I have swabbed with two alcohol swabs, in other words, swabbed it twice), and quickly place the clean stopper and airline into the new jar.
 
I was able to continuously culture T-Iso for appx. one year, and have continuously cultured C-Iso for almost 10 months now, with this method.  I usually split the culture as soon as it gets a dark chocolate brown color, which usually takes around 10 days.  Lately, I've been lazy, and have not needed as much Iso, and so, sometimes I let a culture go as long as 21-28 days before I split it (!).  I guess I've been lucky.  If I let the cultures go for too long, they (1) turn a funny orange color, instead of the golden yellow to dark chocolate brown, and then (2) crash, with most of the cells settling out, and the remainder of the culture turning a funny pinkish orange color.
 
I believe that the key to my success is the use of a microwave to sterilize my media and jars.  The foam stoppers do a good job of sealing the mouth and around the airline, but do allow the air to escape, which may also help vs. a simple hole drilled in a soda bottle cap.  I do currently have a good air filter on my air source, but didn't with my earlier station, and still had good success.
 

Attached Image(s)

JimWelsh
  • Total Posts : 1426
  • Reward points : 1486
  • Joined: 1/22/2010
  • Location: Angwin, CA, US
Re:Culture Journal, Species: Isochrysis galbana - Sunday, August 14, 2011 9:05 PM
Quote Originally Posted by chuenwe


Sweet!  I gotta buy one from SeahourseSource to try again.  I failed miserably using the dish from florida farm.
What pump is that?


The pump is a Whisper AP 300.  I don't know of ANYBODY who has had any success with Isochrysis from FAF disks.  I've started Nannochloropsis and Tetraselmis from FAF disks, but every attempt at Isochrysis from FAF disks has failed for me.  A good starter, like the ones you get from SeahorseSource (via Algagen), is the only way to go for Isochrysis.
 
NOTE:  The pump I currently use is a Whisper AP 300.  The pump in the picture is an AZOO Battery Backup pump.

chuenwe
  • Total Posts : 97
  • Reward points : 113
  • Joined: 7/9/2011
  • Location: Huntington Sta, NY, US
Re:Culture Journal, Species: Isochrysis galbana - Sunday, August 14, 2011 9:16 PM
Should I use C-Iso or T-Iso?  I remember one of them is easier than the other?  
 

JimWelsh
  • Total Posts : 1426
  • Reward points : 1486
  • Joined: 1/22/2010
  • Location: Angwin, CA, US
Re:Culture Journal, Species: Isochrysis galbana - Sunday, August 14, 2011 9:31 PM
I used to use T-Iso.  It worked well for me.  I decided to get C-Iso last year, after I built the new phyto station, only because the SeahorseSource site said it tolerates a wider temperature range, and I was worried about the new phyto station running too warm.  I don't know if it really matters.  The C-Iso has performed very well for me, and seems to survive my neglect (like letting it go as much as 28 days before splitting!) better than the T-Iso did.  Just my $0.02.

chuenwe
  • Total Posts : 97
  • Reward points : 113
  • Joined: 7/9/2011
  • Location: Huntington Sta, NY, US
Re:Culture Journal, Species: Isochrysis galbana - Sunday, August 14, 2011 9:45 PM
Nice!  I'm gonna order a C-Iso to try then.  Thank you!
 

CaptCrash
  • Total Posts : 649
  • Reward points : 1273
  • Joined: 10/23/2011
  • Location: Perth, WA, AU
Re:Culture Journal, Species: Isochrysis galbana - Tuesday, March 27, 2012 7:10 AM
Quote Originally Posted by JimWelsh

Culture medium description:  1200 ml of recycled tank water + 300 ml of fresh RO water microwaved to 85 C (185 F), plus 7.5 ml of FAF Micro Algae Grow (sufficient to make a Guillard F/2 solution for 2000 ml total volume). 
 
 
Jim, is this an error on the amount of FAF Micro Algae Grow?
The bottle lists 30ml to 78 litres, shouldn't you be adding 0.7ml?
It seems like this is way to much Algae Grow (or Im not adding anywhere near enough).

CaptCrash
  • Total Posts : 649
  • Reward points : 1273
  • Joined: 10/23/2011
  • Location: Perth, WA, AU
Re:Culture Journal, Species: Isochrysis galbana - Tuesday, March 27, 2012 7:11 AM
double post

KathyL
  • Total Posts : 2639
  • Reward points : 1504
  • Joined: 6/6/2010
  • Location: St. Louis, MO, US
Re:Culture Journal, Species: Isochrysis galbana - Tuesday, March 27, 2012 7:22 AM
In post #3 Jim uses 0.75ml, but in the top post he says 7 ml. I think its a typo.  0.75 is about right.

Umm_fish?
  • Total Posts : 2835
  • Reward points : 953
  • Joined: 11/4/2009
  • Location: Boulder, CO, US
Re:Culture Journal, Species: Isochrysis galbana - Tuesday, March 27, 2012 9:03 AM
Good job, Jim! I wish I had your perseverance.
--Andy, the bucket man.
"Not to know the mandolin is to argue oneself unknown...." --Clara Lanza, 1886

JimWelsh
  • Total Posts : 1426
  • Reward points : 1486
  • Joined: 1/22/2010
  • Location: Angwin, CA, US
Re:Culture Journal, Species: Isochrysis galbana - Tuesday, March 27, 2012 10:06 AM
Quote Originally Posted by CaptCrash


Quote Originally Posted by JimWelsh

Culture medium description:  1200 ml of recycled tank water + 300 ml of fresh RO water microwaved to 85 C (185 F), plus 7.5 ml of FAF Micro Algae Grow (sufficient to make a Guillard F/2 solution for 2000 ml total volume). 
 

Jim, is this an error on the amount of FAF Micro Algae Grow?
The bottle lists 30ml to 78 litres, shouldn't you be adding 0.7ml?
It seems like this is way to much Algae Grow (or Im not adding anywhere near enough).

 
You and Kathy are correct.  That was a typo that I have now fixed -- thanks for catching it!!!!  The correct amount is 0.75 ml per 2000 ml of media.  Damned decimal points.

CaptCrash
  • Total Posts : 649
  • Reward points : 1273
  • Joined: 10/23/2011
  • Location: Perth, WA, AU
Re:Culture Journal, Species: Isochrysis galbana - Tuesday, March 27, 2012 10:46 AM
Thanks everyone.
 
Im just a bit paranoid Im going to kill my new expensive T-ISO

JimWelsh
  • Total Posts : 1426
  • Reward points : 1486
  • Joined: 1/22/2010
  • Location: Angwin, CA, US
Re:Culture Journal, Species: Isochrysis galbana - Tuesday, March 27, 2012 10:55 AM
Oh, and then it would be all *MY* fault, wouldn't it? 
 
Just kidding -- that was an important typo to catch.  Glad you were paying attention!
 
 

CaptCrash
  • Total Posts : 649
  • Reward points : 1273
  • Joined: 10/23/2011
  • Location: Perth, WA, AU
Re:Culture Journal, Species: Isochrysis galbana - Tuesday, March 27, 2012 10:57 AM
Quote Originally Posted by JimWelsh

 Oh, and then it would be all *MY* fault, wouldn't it? 
 
 
Yup 

JimWelsh
  • Total Posts : 1426
  • Reward points : 1486
  • Joined: 1/22/2010
  • Location: Angwin, CA, US
Re:Culture Journal, Species: Isochrysis galbana - Tuesday, January 22, 2013 1:13 AM
Update:  It seems that almost every year, right about in the October - November period of time, my Isochrysis cultures crash.  This year, I confess, the crash was likely due, at least in part, to me having neglected to do the splits as frequently as I should.  That said, my experience from the past is that another significant contributing factor is the deterioration of the quality of the light due to degredation of the light bulbs over time.  I find that I *MUST* replace the light bulbs at least annually, and based on my experience, they should probably be replaced more frequently than that -- more like every 9 months or so.
 
I've also been re-thinking three aspects of my Isochrysis culture.
 
First:  I think that, for consistency's sake, and to create a more predictable culture, based more on a scientific approach, I'm considering switching from using recycled tank water (with its higher nitrate, phosphate, etc. levels) to using freshly made artificial salt water for my media.
 
Second:  I am also considering perfoming my splits earlier, before the cultures turn a dark brown color.  The reasoning behind this is twofold:  A) To perform the splits while the culture is still actively in the Exponential Growth phase, with a goal of having the subsequent cultures grow more rapidly, and B) Being able to produce a greater total amount of Isochrysis, in terms of absolute cell counts, due to the effects of (A) above.
 
Third:  I am considering increasing the dilution factor of my splits, from the current (VERY conservative) 4:1 ratio, to something more closely approaching 10:1.  I will probably make this change slowly.  In combination with the "Second" adjustment described above, this should increase the overall amount of Isochrysis available to feed my zooplankton without adversely affecting the health of my cultures.

Helix
  • Total Posts : 51
  • Reward points : 87
  • Joined: 3/18/2012
  • Location: Boulder, CO, US
Re:Culture Journal, Species: Isochrysis galbana - Tuesday, January 22, 2013 11:54 AM
Hey Jim,
 
Do you use air filters at all (if so, can you recommend any)?
 
How do you sterilize your nutrients? I notice the culture water becoming cloudy after microwave sterilizing. I presume that some of the nutrients are precipitating (presumably phosphates) when I add the nutrients before microwave sterilizing the media. 
 
 
-Sean
 
BTW, if you need a fresh culture, give me a shout.

JimWelsh
  • Total Posts : 1426
  • Reward points : 1486
  • Joined: 1/22/2010
  • Location: Angwin, CA, US
Re:Culture Journal, Species: Isochrysis galbana - Tuesday, January 22, 2013 12:26 PM
I do use an air filter.  It is basically just a section of 3/4" PVC about 3" long packed really tightly with bulk cotton, together with the fittings necessary to accept the dual airlines coming from my air pump.
 
I do add the Micro Algae Grow before I microwave the media, and I don't have any problem with precipitates.  Conventional wisdom says that the F/2 concentrate is acidic enough to prevent any contaminates from being able to live in it, but I started adding the nutrients before microwaving over a year ago when I was having issues sustaining my cultures.  Like I said above, I'm pretty sure the "root cause" of the problem was having old light bulbs, and not when I added the nutrients, but I've still been microwaving the Micro Algae Grow ever since.
 
The crash was a couple of months ago, and I got new starters a while back, but thanks for the offer!
 

ryannterror
  • Total Posts : 18
  • Reward points : 130
  • Joined: 5/22/2014
  • Location: Fairless hills, PA, US
Re:Culture Journal, Species: Isochrysis galbana - Tuesday, June 24, 2014 12:09 PM
Quote Originally Posted by JimWelsh


Here is a picture of the C-Iso culture today.  I just split the dark culture into two new cultures this morning.  Following my standard phyto protocol, I took 1200 ml of used tank water, plus 300 ml of fresh RO water, in each media jar, plugged the tops with a foam stopper, and microwaved two such media jars in an 1100 watt microwave for 25:80  (Just pushed the keys from the top down the middle column on the keypad, = 26:20).  This amount of time brings the temp up to about 200 F.  It really just needs to get over 185 F, but I like to be certain I've killed everything.  I let it cool to room temperature, and then add roughly 0.75 ml of FAF Micro Algae Grow, which approximates F/2 strength medium.  I then remove the stopper from a fresh media jar, remove the stopper and airline from an existing culture that is dark chocolate brown, and quickly pour about 500 ml from the dark culture into the fresh media.  I then take a prepared clean stopper and airline (either entirely new, or else a used airline I have swabbed with two alcohol swabs, in other words, swabbed it twice), and quickly place the clean stopper and airline into the new jar.

I was able to continuously culture T-Iso for appx. one year, and have continuously cultured C-Iso for almost 10 months now, with this method.  I usually split the culture as soon as it gets a dark chocolate brown color, which usually takes around 10 days.  Lately, I've been lazy, and have not needed as much Iso, and so, sometimes I let a culture go as long as 21-28 days before I split it (!).  I guess I've been lucky.  If I let the cultures go for too long, they (1) turn a funny orange color, instead of the golden yellow to dark chocolate brown, and then (2) crash, with most of the cells settling out, and the remainder of the culture turning a funny pinkish orange color.

I believe that the key to my success is the use of a microwave to sterilize my media and jars.  The foam stoppers do a good job of sealing the mouth and around the airline, but do allow the air to escape, which may also help vs. a simple hole drilled in a soda bottle cap.  I do currently have a good air filter on my air source, but didn't with my earlier station, and still had good success.



 
Jim
 
Where did you get the foam plugs or what type of foam did you use to make them?

picci
  • Total Posts : 4
  • Reward points : 108
  • Joined: 9/20/2013
  • Location: terno d'isola, Bergamo, IT
Re:Culture Journal, Species: Isochrysis galbana - Monday, March 16, 2015 6:02 PM
Hi Jim,
how do you spill the culture when small amount are required? For example few ml for parvo.
 
Cristina
PS I also failed with petri dish.
<message edited by picci on Tuesday, March 24, 2015 5:12 PM>