1. The spectrum of PIG-A gene mutations in aplastic anemia/paroxysmal nocturnal hemoglobinuria (AA/PNH): a high incidence of multiple mutations and evidence of a mutational hot spot
by Yousef Mortazavi, Bruno Merk, Jenny McIntosh, Judith C. W. Marsh, Hubert Schrezenmeier, and Tim R. Rutherford
Blood
Volume 101(7):
April 1, 2003
©2003 by American Society of Hematology

2. PCR strategy for SSCP analysis of PIG-Acoding sequences.
Yousef Mortazavi et al. Blood 2003;101:
©2003 by American Society of Hematology

3. Enrichment of CD59− granulocytes with immunomagnetic beads
Enrichment of CD59− granulocytes with immunomagnetic beads.(A) Fluorescence histogram of CD59 staining of granulocytes before separation, showing a minor negative population.
Enrichment of CD59− granulocytes with immunomagnetic beads.(A) Fluorescence histogram of CD59 staining of granulocytes before separation, showing a minor negative population. (B) Fluorescence histogram after depletion of CD59+ cells.
Yousef Mortazavi et al. Blood 2003;101:
©2003 by American Society of Hematology

4. DNA sequence derived from a heteroduplex fragment
DNA sequence derived from a heteroduplex fragment.A heteroduplex band from patient no. 34 was isolated, reamplified, and sequenced.
DNA sequence derived from a heteroduplex fragment.A heteroduplex band from patient no. 34 was isolated, reamplified, and sequenced. Note unambiguous sequence up to the arrowed nucleotide (T at nt 775 on the PIG-A cDNA sequence), then ambiguous sequence. By subtracting the normal sequence 775-GTCAGCAGACTTGTT from the observed sequence the alternative peaks can be clearly read as 778-AGCAGACTTGTTAC showing deletion of 3 nucleotides.
Yousef Mortazavi et al. Blood 2003;101:
©2003 by American Society of Hematology

5. DNA sequence showing a 31-nt duplication
DNA sequence showing a 31-nt duplication.An abnormal band from patient no. 24 was isolated, reamplified, and sequenced.
DNA sequence showing a 31-nt duplication.An abnormal band from patient no. 24 was isolated, reamplified, and sequenced. The duplicated sequence is shown overlined.
Yousef Mortazavi et al. Blood 2003;101:
©2003 by American Society of Hematology

6. Mutation-specific PCR products
Mutation-specific PCR products.Patient DNA was amplified with primers specific for the 5-bp deletion at nt 662 to 666.
Mutation-specific PCR products.Patient DNA was amplified with primers specific for the 5-bp deletion at nt 662 to 666. The identity of the bands at 163 bp was confirmed by DNA sequencing. M indicates size marker.
Yousef Mortazavi et al. Blood 2003;101:
©2003 by American Society of Hematology

7. PNH Type I, II, and III populations in granulocytes
PNH Type I, II, and III populations in granulocytes.Fluorescence histograms of CD59-stained granulocytes from patients (A) no. 22, (B) and (C) no. 23, (D) no. 25, (E) and (F) no. 28, (G) and (H) no. 29, before (A-B,D-E,G) and after (C,F,H) immunomagnetic de...
PNH Type I, II, and III populations in granulocytes.Fluorescence histograms of CD59-stained granulocytes from patients (A) no. 22, (B) and (C) no. 23, (D) no. 25, (E) and (F) no. 28, (G) and (H) no. 29, before (A-B,D-E,G) and after (C,F,H) immunomagnetic depletion of CD59+ cells. Unfilled histogram indicates healthy control; filled histogram, patient.
Yousef Mortazavi et al. Blood 2003;101:
©2003 by American Society of Hematology