Abstract
Flow cytometric methods are commonly used to analyze white blood cell surface antigen expression. We developed a flow cytometric method to detect red blood cell (RBC)-bound immunoglobulin (Ig)G, IgA, and IgM. RBCs were washed; incubated with fluorescein isothiocyanate (FITC)-conjugated anti-IgG, -IgA, or -IgM; washed; and analyzed on the flow cytometer. The method was optimized by determining the dilution of FITC-conjugated anti-IgG, -IgA, and -IgM providing the greatest amount of fluorescence when tested with Ig-coated RBCs and the least amount of fluorescence when tested with naive RBCs. Tannic acid was used to prepare Ig-coated RBCs. Cross-reactivity of FITC-conjugated anti-IgG, -IgA, and -IgM with Ig-coated RBCs was evaluated, and a reference range was established. Use of this method may assist in clinical evaluation of patients who present with hemolysis and a negative direct antiglobulin test.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 161-169 |
| Number of pages | 9 |
| Journal | Immunohematology |
| Volume | 32 |
| Issue number | 4 |
| State | Published - Jan 1 2016 |
| Externally published | Yes |
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Keywords
- Autoimmune hemolytic anemia
- Direct antiglobulin test
- Flow cytometry
- Immunoglobulins
- Red blood cell
- Tannic acid
ASJC Scopus subject areas
- Medicine(all)
Cite this
A detailed flow cytometric method for detection of low-level in vivo red blood cell-bound IgG, IgA, and IgM. / Beres, Wendy; Meny, Geralyn M.; Nance, Sandra.
In: Immunohematology, Vol. 32, No. 4, 01.01.2016, p. 161-169.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - A detailed flow cytometric method for detection of low-level in vivo red blood cell-bound IgG, IgA, and IgM
AU - Beres, Wendy
AU - Meny, Geralyn M.
AU - Nance, Sandra
PY - 2016/1/1
Y1 - 2016/1/1
N2 - Flow cytometric methods are commonly used to analyze white blood cell surface antigen expression. We developed a flow cytometric method to detect red blood cell (RBC)-bound immunoglobulin (Ig)G, IgA, and IgM. RBCs were washed; incubated with fluorescein isothiocyanate (FITC)-conjugated anti-IgG, -IgA, or -IgM; washed; and analyzed on the flow cytometer. The method was optimized by determining the dilution of FITC-conjugated anti-IgG, -IgA, and -IgM providing the greatest amount of fluorescence when tested with Ig-coated RBCs and the least amount of fluorescence when tested with naive RBCs. Tannic acid was used to prepare Ig-coated RBCs. Cross-reactivity of FITC-conjugated anti-IgG, -IgA, and -IgM with Ig-coated RBCs was evaluated, and a reference range was established. Use of this method may assist in clinical evaluation of patients who present with hemolysis and a negative direct antiglobulin test.
AB - Flow cytometric methods are commonly used to analyze white blood cell surface antigen expression. We developed a flow cytometric method to detect red blood cell (RBC)-bound immunoglobulin (Ig)G, IgA, and IgM. RBCs were washed; incubated with fluorescein isothiocyanate (FITC)-conjugated anti-IgG, -IgA, or -IgM; washed; and analyzed on the flow cytometer. The method was optimized by determining the dilution of FITC-conjugated anti-IgG, -IgA, and -IgM providing the greatest amount of fluorescence when tested with Ig-coated RBCs and the least amount of fluorescence when tested with naive RBCs. Tannic acid was used to prepare Ig-coated RBCs. Cross-reactivity of FITC-conjugated anti-IgG, -IgA, and -IgM with Ig-coated RBCs was evaluated, and a reference range was established. Use of this method may assist in clinical evaluation of patients who present with hemolysis and a negative direct antiglobulin test.
KW - Autoimmune hemolytic anemia
KW - Direct antiglobulin test
KW - Flow cytometry
KW - Immunoglobulins
KW - Red blood cell
KW - Tannic acid
UR - http://www.scopus.com/inward/record.url?scp=85028503314&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85028503314&partnerID=8YFLogxK
M3 - Article
C2 - 28257230
AN - SCOPUS:85028503314
VL - 32
SP - 161
EP - 169
JO - Immunohematology / American Red Cross
JF - Immunohematology / American Red Cross
SN - 0894-203X
IS - 4
ER -