Application - SPEC114220010008

DOE-SPEC-1142-2001

that have been identified by positive blood proliferation responses to beryllium have

gone on to develop disease.

4. APPLICATION. Beryllium sensitivity testing is used as a screening tool for possible chronic

beryllium disease, as a surveillance tool in indicating hazardous working conditions, and as

part of the diagnostic criteria for the disease. The blood lymphocyte proliferation test for

beryllium sensitization (BeLPT) is a screening test with a sensitivity and specificity that is

not clearly defined at this time. This is attributable to the fact that the populations of normal

people have not had lung biopsy and bronchoalveolar lavage to include or exclude chronic

beryllium disease. The value of a repeatedly positive BeLPTs in predicting CBD can be

calculated and is estimated to be approximately 44 to 50 percent (positive predictive value).

The bronchoalveolar lavage lymphocyte proliferation test for beryllium sensitization (BAL-

LPT) is the preferred test of beryllium sensitivity as part of the diagnostic criteria for chronic

beryllium disease.

5. TRITIATED THYMIDINE UPTAKE PROCEDURE FOR DETERMINING BERYLLIUM

SENSITIZATION

5.1.

REAGENTS AND EQUIPMENT

5.1.1. RPMI-1640 medium supplemented with 25mM HEPES buffer and 200 mM

L-glutamine

5.1.2. Human serum, Type AB, heat inactivated to eliminate complement activity.

Each lot MUST BE TESTED for its ability to support lymphocyte

proliferation (i.e., it should not exhibit excessive levels of toxicity or cell

killing in the presence of beryllium). It is aliquoted and stored at -20C until

use. It should not be refrozen.

5.1.3. Penicillin-streptomycin, 100 U and 50 gm/ml respectively or 50 g/ml

gentamicin sulfate

5.1.4. Phosphate-buffered balanced salt solution (i.e., Hank's, Dulbecco's, etc).

5.1.5. Tritiated Thymidine (specific activity 2-10 Ci/mM)

5.1.6. Beryllium sulfate, 0.2 M solution (Brush Wellman, Inc).

5.1.7. PHA (Phytohemagglutinim) and/or Con A (concanavalin-A) and/or Candida

albicans allergenic extract or recall antigen suitable for population such as

tetanus. Should be prepared in a concentration known to stimulate lymphocyte

proliferation optimally (i.e., ~30 g/ml in culture and 10 g/ml in culture for

PHA and ConA, respectively).