SOP for ABO Plasma Grouping in Gel requires 50ul of plasma. When unexpected agglutination (suspected presence of rouleaux) occurs performing ABO Plasma Grouping in Gel, would it be useful to repeat the test in Gel using 25uL of plasma and 25uL of 0.9% buffered saline as an investigative tool?

NO!  The whole point of column agglutination technology is that the ionic strength has to be the same, otherwise you will end up with either false positives or false negatives (see the inserts).

19 hours ago, Malcolm Needs said:

NO!  The whole point of column agglutination technology is that the ionic strength has to be the same, otherwise you will end up with either false positives or false negatives (see the inserts).

How does the ionic-strength of 50uL of plasma differ from 50uL (25uL of plasma + 25uL of Buffered Saline)?

I understand the importance of maintaining a low-ionic strength environment for indirect antiglobulin testing at 37C in Anti-IgG Gel, but not in direct agglutination tests at room temperature in Buffered Gel.  Historically, low-ionic strength test systems were not implemented in direct agglutination test systems. 

Please elaborate.

58 minutes ago, Dansket said:

How does the ionic-strength of 50uL of plasma differ from 50uL (25uL of plasma + 25uL of Buffered Saline)?

I understand the importance of maintaining a low-ionic strength environment for indirect antiglobulin testing at 37C in Anti-IgG Gel, but not in direct agglutination tests at room temperature in Buffered Gel.  Historically, low-ionic strength test systems were not implemented in direct agglutination test systems. 

Please elaborate.

I am no chemist Dansket!  I am hoping that Anna Galvania will be able to answer this, as she used to work for DiaMed in Switzerland, and knows MUCH more about this than do I.

4 hours ago, Malcolm Needs said:

I am no chemist Dansket!  I am hoping that Anna Galvania will be able to answer this, as she used to work for DiaMed in Switzerland, and knows MUCH more about this than do I.

Wanna learn a site trick?  If you type the @ symbol followed by their user name, the user usually gets an email notification with a link to this thread.  Like this:: @galvania.

@galvania

Sorry, have only just seen this.  Now that I am retired >I no longer check my e-mails every day. (yes, that does say day, not hour.  I know the youngsters will find that totally unbelievable).

Anyway, if you suspect the presence of rouleaux it can be very difficult to differentiate between rouleaux and agglutination in gel although rouleaux does tend to have a pink haze about it.  Even more difficult if BOTH are present….

I would not try to elucidate this in gel.  I would do this in a tube and look at the result down a microscope (one of the few times where I think use of a microscope is justified) and then you can visually see the difference.

As for the other part of the question - normal saline can interfere with the buffers used in the gel itself - regardless of the technique used (IAT, direct agglutination etc).  This can lead to unagglutinated cells 'hanging' in the gel and this gives a rosy appearance which can be quite strong.  It does not always happen but when it does can be mistaken for a positive result.  A negative result can be interpreted as a true negative - saline will not weaken a result in a direct agglutination test.  

 

As an aside to Cliff, I didn't get an email notification but that could be because I have not updated my e-mail address!!!!!  Will do so as soon as posible.  Have changed countries too.  Am now in Italy

Cliff - I cant see where to edit my e-mail address.  It is now <removed by admin>.  And no - I have no problem that anyone in the forum can see this

Edited January 5, 20196 yr by Cliff
Removed email address

23 hours ago, galvania said:

Cliff - I cant see where to edit my e-mail address.  It is now <removed by admin>.  And no - I have no problem that anyone in the forum can see this

I have updated your email for you.  I appreciate that you shared your email with everyone on the site, but the whole world could see it.  There are lots of bots that roam around trying to find email addresses to send you spam.  I have removed it from the post.