Dataset Information

DEA 1 expression on dog erythrocytes analyzed by immunochromatographic and flow cytometric techniques.

ABSTRACT:

Background

The Dog erythrocyte antigen (DEA) 1 blood group system was thought to contain types DEA 1.1 and 1.2 (and possibly 1.3 [A3]). However, DEA 1.2+ dogs are very rare and newer typing methods reveal varying degrees of DEA 1 positivity.

Objectives

To assess if variation in DEA 1 positivity is because of quantitative differences in surface antigen expression. To determine expression patterns in dogs over time and effects of blood storage (4°C). To evaluate DEA 1.2+ samples by DEA 1 typing methods.

Animals

Anticoagulated blood samples from 66 dogs in a research colony and from a hospital, and 9 previously typed DEA 1.2+ dogs from an animal blood bank.

Methods

Research study: Samples were analyzed by flow cytometry and immunochromatographic strip using a monoclonal anti-DEA 1 antibody.

Results

Twenty dogs were DEA 1-, whereas 46 dogs were weakly to strongly DEA 1+. Antigen quantification revealed excellent correlation between strip and flow cytometry (r = 0.929). Both methods reclassified DEA 1.2+ samples as weakly to moderately DEA 1+, but they were not retyped with the polyclonal anti-DEA 1.1/1.X antibodies. Dogs and blood samples retained their relative DEA 1 antigen densities over time.

Conclusions and clinical importance

The blood group system DEA 1 is a continuum from negative to strongly positive antigen expression. Previously typed DEA 1.2+ appears to be DEA 1+. These findings further the understanding of the DEA 1 system and suggest that all alleles within the DEA 1 system have a similarly based epitope recognized by the monoclonal antibody.

SUBMITTER: Acierno MM

PROVIDER: S-EPMC3998747 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:BACKGROUND: Malaria pigment (haemozoin, Hz) has been the focus of diverse research efforts. However, identification of Hz-containing leukocytes or parasitized erythrocytes is usually based on microscopy, with inherent limitations. Flow cytometric detection of depolarized Side-Scatter is more accurate and its adaptation to common bench top flow cytometers might allow several applications. These can range from the ex-vivo and in-vitro detection and functional analysis of Hz-containing leukocytes to the detection of parasitized Red-Blood-Cells (pRBCs) to assess antimalarial activity. METHODS: A standard benchtop flow cytometer was adapted to detect depolarized Side-Scatter. Synthetic and Plasmodium falciparum Hz were incubated with whole blood and PBMCs to detect Hz-containing leukocytes and CD16 expression on monocytes. C5BL/6 mice were infected with Plasmodium berghei ANKA or P. berghei NK65 and Hz-containing leukocytes were analysed using CD11b and Gr1 expression. Parasitized RBC from infected mice were identified using anti-Ter119 and SYBR green I and were analysed for depolarized Side Scatter. A highly depolarizing RBC population was monitored in an in-vitro culture incubated with chloroquine or quinine. RESULTS: A flow cytometer can be easily adapted to detect depolarized Side-Scatter and thus, intracellular Hz. The detection and counting of Hz containing leukocytes in fresh human or mouse blood, as well as in leukocytes from in-vitro experiments was rapid and easy. Analysis of CD14/CD16 and CD11b/Gr1 monocyte expression in human or mouse blood, in a mixed populations of Hz-containing and non-containing monocytes, appears to show distinct patterns in both types of cells. Hz-containing pRBC and different maturation stages could be detected in blood from infected mice. The analysis of a highly depolarizing population that contained mature pRBC allowed to assess the effect of chloroquine and quinine after only 2 and 4 hours, respectively. CONCLUSIONS: A simple modification of a flow cytometer allows for rapid and reliable detection and quantification of Hz-containing leukocytes and the analysis of differential surface marker expression in the same sample of Hz-containing versus non-Hz-containing leukocytes. Importantly, it distinguishes different maturation stages of parasitized RBC and may be the basis of a rapid no-added-reagent drug sensitivity assay.