Trypsin is a valuable enzyme used in our IRL.  In order to determine if treatment is successful, we test trypsin treated reagent red cells with N lectin (Vicea Graminea).  Trypsin treated N negative red cells test positive with N lectin.  Unfortunately, we have been informed that our current suplier of N lectin is stopping production.  We are in search of a new source of N lectin and are wondering what other labs are using.  We have procedures for making the lectin ourselves, but we are unable to find a supplier of Vicea Graminea seeds.  Any information is appreciated. 

I purchase anti-N lectin from HemoBioscience

I purchase anti-N lectin from HemoBioscience

We currently purchase from them as well.  We were informed that they are no longer going to be producing/selling N lectin.

Why don't you just test treated and untreated cells in parallel with a normal anti-M or anti-N?  The untreated cells will be positive and the trypsin treated cells will be negative.  Won't they??

Why don't you just test treated and untreated cells in parallel with a normal anti-M or anti-N?  The untreated cells will be positive and the trypsin treated cells will be negative.  Won't they??

Thanks for your response!  I agree that this seems like a logical "check" of treatment and we tried this with Anti-M and the cells were still weakly positive.  I don’t believe that the antigens are always destroyed, but are “altered.”  We did not feel like this was a good test of treatment as it could be very subjective and not as clear cut as N negative becomes macroscopically positive.  While the literature I read does state that the M and N antigens are denatured, this was not a suggested method of checking the enzyme treatment.  The published method is to test treated N negative cells with N lectin.  Since the publication tell us to use N lectin, we were hoping to find another source of N lectin so that we would not have to complete a huge validation on another method that is not as reliable. 

I use anti-M (monoclonal from Immucor) to test my papain pretreatment.  Works great.  I test in gel.  I did pretreat an anti-N last week with excellent results (using lectin).  I couldn't find another N lectin - only anti-M or anti-N.  I believe enzyme cleaves portions of GPA that contain M/N ags. I have never seen a study that says these ags are altered by enzyme.  It is also the reason Ss are variable with enzyme as GPB gets cleaved right around the area of those ags (and U is enhanced because that ag on GPB is very close to the red cell surface and is therefore more available to ab).

 

I would think that if you got a significant reduction in strength, even though not completely negative, that this would indicate a successful enzyme pretreatment.  However, it is up to you as to how you interpret those results. 

I found this on the internet. Please let us know the outcome if you look into this.   

 

http://eylabs.com/product/anti-n-vicia-graminea-lectin-vga-2ml/?distributor=true#.VlUmFexVhBc

You could consider using Glycine soja for your control. Works well for us. I actually have some vicia graminea seeds here but we have never had much luck with making a successful lectin with them.