Phosphate buffered saline versus saline for blood bank laboratory testing

A colleague in South Carolina comments that some of their blood bank laboratory reagents have new direction circulars that read "use phosphate buffered saline" (PBS). They have always used certified saline solutions at her facility. As far as she knows, the difference is in pH. The circular for the saline that they use gives a pH range of 6.0-7.5. She thinks PBS has a pH of 7.4. Her question is should they change the saline that they are using? If controls are working, shouldn't it be OK to use the saline that they have used for years?

The following comments have been received in response.

ADDENDA Dec. 16, 2010

1. A medical technologist at a community hospital in Iowa writes "We switched to phosphate buffered saline after noticing a change in a package insert. Our controls always properly worked with 'blood bank saline' at a different pH." He states that the way it read required the change. Their validation "consisted of parallel testing control and various post partum patient specimens with the old ('blood bank saline') and new (PBS) salines. Two of the patient specimens consisted of moms' bloods 'spiked' with a borderline amount of cord cells to produce weakly positive specimens." He reports no apparent difference in results obtained between the two systems.

ADDENDA Dec. 23, 2010

2. According to W. John Judd, FIBMS, MIBiol, Emeritus Professor of Immunohematology at the University of Michigan (attribution used with permission), along with certified saline (pH 6.5-8.0), standard blood bank saline is available commercially at pH 6.5-7.0. In his opinion, none of these products can be considered to be buffered saline, which is why there are three distinct types of saline listed in reagent catalogues. According to Professor Judd, it seems that buffered saline at pH 7.2 is marketed to "enhance weak serological reactions." He wonders what testing was done to convince the FDA to allow such a claim! Buffered saline at pH 7.3 * 0.1 can also be prepared in-house, using the formula in Methods in Immunohematology, which is available from the AABB Bookstore. Does it make a difference if you don't use buffered saline? When performing ABO and Rh typing, the antisera are buffered anyway, and apart from one anti-B product no longer available, Professor Judd has never encountered RBCs ABO typing problems related to pH. For reverse ABO typing, the test plasma or serum maintain the test milieu close to a neutral pH. Further, some antibodies such as anti-M give enhanced reactions at a slightly acidic pH (e.g. 6.2). When performing antiglobulin tests, saline that is too acidic may cause antibody to dissociate during washing, thereby yielding unwanted weak or negative tests. Professor Judd wishes reagent manufacturers would not place such restrictions on the use of their products, but they tested their products that way when developing data for the FDA. Colleagues in the field need to comply with the product circular unless meaningful data can be provided for an alternative method. One could argue that past testing results do not demonstrate equivalency since it is unknown how many unwanted negatives were encountered in the past.

ADDENDA Jan. 20, 2011

3. Dr. George Garratty and Patricia A. Arndt, MS, MT(ASCP)SBB, Research Department, American Red Cross Blood Services (attribution used with permission) report that in the 1960s, data were published (e.g., Hughes-Jones et al. Immunology 1964;7:72-81) showing that Rh antibodies reacted optimally at pH 6.5 to 7, although there was little difference over the pH range of 5.5 to 8.5. They write: "Following this, many blood bank laboratories (including their own) started using PBS to optimize detection of weak antibodies. As usual, there are exceptions, e.g., some anti-M prefer low pH. Specific commercial saline products have become available (e.g., at a suitable pH) for use in immunohematology (see Tables 1 and 2 for some examples). Pharmaceutical saline may have a lower pH and may not be suitable for immunohematology. PBS can be prepared by mixing different proportions of acidic and alkaline stock solutions to reach a certain pH (see AABB Technical Manual, 16th ed., p 895). Your routine controls would not be suitable to control for this pH optimization, as only certain antibodies would be affected by pH. The best QC would be to assure that your saline is within the suitable pH range.
   
   We should all note that the recommendations above, and possibly the development of special saline for blood banks, and the recommendations in the package inserts were based on old (excellent) data concerning antigen-antibody reactions using saline-suspended RBCs. Although probably still relevant to Reference Labs, it may not be so relevant nowadays for hospitals, with increased use of the gel test and solid phase automated methods. We know of no data on the effect of pH in these approaches; nor do we know of data on the effect of pH on interactions of anti-IgG with IgG-coated RBCs [the major use of saline (before the gel test) was washing RBCs for the antiglobulin test]. Regardless of this, unfortunately if a specific type of saline solution is mentioned in a package insert, then one has to follow those instructions".

Table 1: Some unbuffered blood bank saline products:

Manufacturer and/or Distributor	Product Name (#)	pH
Azer Scientific	Blood Bank Saline Solution (ES-1240, ES-1244)	Between 6.5 and 7.0
Cardinal Health (Thermo Scientific)	SP® Certified Blood Bank Saline (B3158-1,2, or 3)	6.0-7.5
Cole-Parmer	Sodium chloride, 0.9% (w/v) Aqueous Solution, Isotonic Saline, Suitable for Use as Blood Bank Saline (EW-86975-82, EW-86982-66, EW-86987-56, EW-86989-09, EW-86994-06)
Ever Scientific (Cardinal Health)	Blood Bank Saline, standard solution (ES1244)	6.5-7.0
Fisher Scientific	Fisherbrand* Blood Bank Saline (23-293-184, 23-062-125, 23535-435)	6.0-7.5
Thermo Scientific	Blood Bank Saline (23-062-125, 23-293-184, 23-535-435)	6.0-7.5

 

Table 2: Some buffered blood bank saline products:

Manufacturer and/or Distributor	Product Name (#)	pH
Azer Scientific	Blood Bank Saline Solution (ES-1243)	7.0-7.2
Cardinal Health (Thermo Scientific)	Blood Bank Saline, pH 7.0 to 7.2 (B3158-8 or 6)	7.0-7.2
Casco-Nerl Diagnostics	Saline Solution (8504, 8505)	7.0-7.2
EMD Chemicals	Blood Bank Immuno-Saline Solution (EM-R01826-84)	7.0-7.5
EMD Chemicals	Blood Bank Immuno-Saline Plus Solution (EM-R01830-82)	6.7-7.1
Ever Scientific (Cardinal Health)	Blood Bank Saline, buffered solution (ES1243)	7.0-7.2
Fisher Scientific	Fisherbrand* Blood Bank Saline (23-312-651, 23-309-178)	7.2 ± 0.1
Hemobioscience	Phosphate Buffered Saline Concentrate (H307)	7.00 ± 0.05
NERL/Thermo Fisher Scientific	Buffered Blood Bank Saline pH 7.0-7.2 (8504, 8505)	7.0-7.2
Thermo Scientific	Blood Bank Saline, Buffered (23-309-178, 23-312-651)	7.2 ± 0.1

 

Submit comments to the e-Network Forum at enetworkforum@cbbsweb.org

Ira A. Shulman, MD
CBBS e-Network Forum Senior Editor & Moderator

W. Tait Stevens, MD
CBBS e-Network Forum Editor & Moderator

Elizabeth M. St. Lezin, MD
CBBS e-Network Forum Associate Editor & Moderator