FLOATING LEAF DISC LAB .k12.sd.us



FLOATING LEAF DISC LAB

In using the "leaf disk assay" lab, has any one had a  problem getting the leaf disks to sink?  Then once we did & started timing----none floated!  This should be a simple activity, getting predictable results.  Any ideas?  We used spinich leaves.

Patty, Infiltration of the leaves requires a drop of detergent in the sodium bicarbonate solution to work. I tried spinach last year and it took almost an hour. I used ivy this year and the disks floated in about eleven minutes.

Patti:

Sounds  like you have an experiment on your hands.  I encourage you to keep on trying this lab because it does work very reliably once you learn a few of the things that throw a wrench into the procedure and it really is a fun lab to do with a lot of possibility for actual student-led research.

You may have already gone to my web page at:



If not, I expect that you will find answers to your questions, there.  I don't really have enough information in your post to do more than speculate but having used this technique for many years and having presented it to hundred's of teachers I can offer some of the common slip-ups.....

Regarding sinking the disks--this is usually the bottleneck in this lab.  You must put the disk under a vacuum to infiltrate the inner leaf with bicarb solution....I use syringes as a vacuum source but have also used aspirators and vacuum pumps.   Too much vacuum or too many attempts at sinking the disks can actually disrupt the cell membranes and damage the disks.  The key to sinking the disks is to overcome the hydrophobic cuticle with very dilute soap solution.  If you use the syringe vacuum technique described on the web page then it shouldn't take more than two vacuum events to sink the disks.  If it does add a few more drops of soap solution and the disks should sink on the next vacuum event.

Regarding no disk rising....

Could be you didn't allow enough time or have enough light.  Spinach usually comes up within 8 minutes if under bright light (within 4 to 6 inches of a bright 30 watt spiral flourescent bulb as shown on the web page).  The most common reason for disks not floating is forgetting to add a bit of sodium bicarbonate as a CO2 source.  This past weekend, at a workshop, only one participant had trouble with sinking his disks and his disks did not rise either.  I suspect that in his case that he damaged the disks while trying to sink them.  The water was green from damaged cells.  If the leaf and cell structure are not in place the procedure will not work. 

Ann, I used Dawn dishwashing detergent at full strength, but only put one or two drops in 500mL of 0.1 % sodium bicarbonate solution.

BLAIR BUCK

Disk Users;

We use a surfactant available at places that use wettable powders in some application or another.  We get ours from our local country club  superintendent. Have also gotten it from a florist.  One bottle will last a career (unless you drop it). Dose = .05  ml in 100 mL of buffer.

Hopefully you are all using 50 mL (or smaller) syringes to infiltrate the disks and not expensive/frustrating vacuum pumps. Remove the syringe barrel, put in the disks (try not to touch or damage-use a disposable bacteria loop, etc), suck up 25 mL infiltration sol'n, put your finger on the needle end and pull back to 49 mL.  Swirl gently. Don't shake!  No bubbles!   Repeat 3x. Disks will sink like stones. Cover syringe in foil or work in very low light until students get enough disks. Transfer to run tubes. Don't forget the bicarbonate. We run 20 disks in a 100 mL flat bottom tube (approx. length = 40 cm.). First kid to get 90% of disks to rise wins a prize (old AP Biol T-shirt, AP review guide used for 1 night by some kid who got a 2 on the test, model molecule kit, etc.).

This is my favorite lab. I could write about it for hours. Original protocol in ABT many years ago.

My AP Biology classes just finished Lab 1 today (we started school 8/28).  Some of them actually think they are enrolled in "AP Potato." Pray for us.

If you want more info. by snail mail send us your address. If you want a digital photo of our set-up respond electronically. It may take us a while.

W. Anderson

Sandra Litvin

Unionville HS

Kennett Square, PA

................
................

In order to avoid copyright disputes, this page is only a partial summary.

Google Online Preview   Download