An ELISA-based procedure for assaying proteins in digests of human leukocytes and cell lines, using specifically selected peptides and appropriate antibodies

Table 2 Summary of presence of P-glycoprotein in patient samples, as detected with the Peptidomatrix and the Rhodamine uptake assay.

	Patient	Type of leukemia	Peptidomatrix results (ng peptide/mg protein)	Peptidomatrix results (Grade)	Status by FACS (Grade)
	CEM wt		7	Negative
	CEM col1000		114	Positive
1	AP	AML	72.5	Positive (low)	Positive
2	DA	T-ALL	26.37	Positive (low)	Negative
3	GB	t-AML	150	Positive (high)	Positive
4	KA	T-ALL	0	Negative	Negative
5	RE	AML	144	Positive (high)	Positive
6	WE	AML	12.2	Positive (low)	Positive (low)
7	SA	AML	7	Negative	Negative
8	SN	AML	90.93	Positive (low)	Positive
9	SY	CLL	209	Positive (high)	Positive
10	WR	APL	4.9	Negative	Negative
11	YN	AML	7.9	Negative	Negative
12	CM	AML	6	Negative	Negative
13	DM	CML	62.27	Positive (low)	Positive
14	GG	AML	4.8	Negative	Negative
15	SG	AML	56.4	Positive (low)	Positive
16	OZ	AML	38.5	Positive (low)	Positive (low)

The table compares the results that we have from the clinical functional method, Rhodamine 123 uptake and the Peptidomatrix assay (see text). The Rhodamine 123 uptake assay is based on the differential dye retention in the absence and presence of Verapamil, an inhibitor of Pgp. The MDR1 protein levels in the samples were determined by the Peptidomatrix using the MDR P3 peptide and are expressed as ng peptide per mg protein. Multidrug sensitive CEM cells (wt) and resistant CEM cells (col1000) served as controls. To allow a comparison with the semiquantitative data of the other assay a gradation of MDR1 levels was defined by comparison to the control cells: The samples that had levels similar to the wt cells ± 10 % (up to 8 ng/mg protein) were graded as negative, the samples that had levels similar to the col1000 cells (100–125 ng/mg) were graded as positive, the samples that had levels higher (> 125 ng/mg) or lower then the col1000 cells (8–100 ng/mg) were graded as high-positive or low-positive respectively. The data shown are averages of duplicates, the difference between the duplicates were below 5%.

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