Hans‐Peter Spengler

Basics of flow cytometry–based sterility testing of platelet concentrates

BACKGROUND:  Flow cytometry (FACS) is a common technique in blood banking. It is used, for example, for the enumeration of residual white blood cells in plasma and in cellular blood products. It was investigated whether it can also be applied for sterility testing of buffy coat–derived platelet concentrates (PCs).

A comparison of three rapid bacterial detection methods under simulated real-life conditions.

BACKGROUND: Bacterial screening of all produced platelet concentrates (PCs) is implemented in many countries to reduce the risk of transfusion‐transmitted sepsis. This study compares three rapid bacterial detection methods by imitating real‐life conditions.

Bacteria detection by flow cytometry

Abstract Since bacterial infection of the recipient has become the most frequent infection risk in transfusion medicine, suitable methods for bacteria detection in blood components are of great interest. Platelet concentrates are currently the focus of attention, as they are stored under temperature conditions, which enable the multiplication of most bacteria species contaminating blood donations. Rapid methods for bacteria detection allow testing immediately before transfusion in a bed-side like manner. This approach would overcome the sampling error observed in early sampling combined with culturing of bacteria and would, at least, prevent the transfusion of highly contaminated blood components leading to acute septic shock or even death of the patient. Flow cytometry has been demonstrated to be a rapid and feasible approach for detection of bacteria in platelet concentrates. The general aim of the current study was to develop protocols for the application of this technique under routine conditions. The effect of improved test reagents on practicability and sensitivity of the method is evaluated. Furthermore, the implementation of fluorescent absolute count beads as an internal standard is demonstrated. A simplified pre-incubation procedure has been undertaken to diminish the detection limit in a pragmatic manner. Additionally, the application of bacteria detection by flow cytometry as a culture method is shown, i.e., transfer of samples from platelet concentrates into a satellite bag, incubation of the latter at 37°C, and measuring the contaminating bacteria in a flow cytometer. Clin Chem Lab Med 2008;46:947–53.

Sterility testing of platelet concentrates prepared from deliberately infected blood donations

BACKGROUND:  In general the bacterial count in freshly donated blood is low and even lower in the corresponding platelet concentrates (PCs). By use of flow cytometry (FACS) for sterility testing, the reliability of early versus later sampling times was evaluated.

Flow cytometric assay for the simultaneous determination of residual white blood cells, red blood cells, and platelets in fresh‐frozen plasma: validation and two years' experience

BACKGROUND: A fully automated single‐tube assay with tubes (BD TruCOUNT, BD Biosciences) for absolute counting of residual cells in freshly prepared plasma by flow cytometry was developed (BD Plasma Count).

BLOOD COMPONENTS: A new one‐platform flow cytometric method for residual cell counting in platelet concentrates

BACKGROUND: According to German regulations and guidelines, residual red blood cells (rRBCs) and residual white blood cells (rWBCs) must number fewer than 3 × 109 cells/unit and 1 × 106 cells/unit in platelet concentrates (PCs), respectively. Due to low levels of residual cells in final products, there is still a need for fast, reliable, and sensitive methods of automated detection of these cell types.

A new one-platform flow cytometric method for residual cell counting in platelet concentrates.

BACKGROUND: According to German regulations and guidelines, residual red blood cells (rRBCs) and residual white blood cells (rWBCs) must number fewer than 3 × 109 cells/unit and 1 × 106 cells/unit in platelet concentrates (PCs), respectively. Due to low levels of residual cells in final products, there is still a need for fast, reliable, and sensitive methods of automated detection of these cell types.

A flow cytometric method for platelet counting in platelet concentrates

BACKGROUND: The platelets (PLTs) in PLT concentrates are counted with hematology analyzers, but varying results among different hematology analyzers are observed, making comparisons very difficult. Due to the absence of red blood cells in PLT concentrates, the International Council for Standardization in Hematology (ICSH) reference method was modified to be used for PLT concentrates and validated in an international comparative study.

BLOOD COMPONENTS: A new one-platform flow cytometric method for residual cell counting in platelet concentrates: RESIDUAL CELL DETERMINATION BY FLOW CYTOMETRIC THROMBO COUNT ASSAY

BACKGROUND: According to German regulations and guidelines, residual red blood cells (rRBCs) and residual white blood cells (rWBCs) must number fewer than 3 × 109 cells/unit and 1 × 106 cells/unit in platelet concentrates (PCs), respectively. Due to low levels of residual cells in final products, there is still a need for fast, reliable, and sensitive methods of automated detection of these cell types.

Improvement of bacteria detection by flow cytometry (BDFC) / Verbesserter Nachweis von Bakterien mittels Durchflusszytometrie

Abstract Since bacterial infection of the recipient has become the most frequent infection risk in transfusion medicine, suitable methods for bacteria detection in blood components are of great interest. Platelet concentrates are currently in the focus of attention as they are stored under temperature conditions, which enable the multiplication of most bacteria species contaminating blood donations. Rapid methods for bacteria detection allow testing immediately before transfusion in a bedside-like manner. This approach would overcome the sampling error observed in early sampling combined with culturing of bacteria and would at least prevent the transfusion of highly contaminated blood components leading to acute septic shock or even death of the patient. Flow cytometry has been demonstrated to be a rapid and feasible approach for the detection of bacteria in platelet concentrates. The general aim of the current study is to develop protocols for the application of this technique under routine conditions. The effect of improved test reagents on practicability and sensitivity of the method is evaluated. Furthermore, the implementation of fluorescent absolute count beads as an internal standard is demonstrated. A simplified pre-incubation procedure has been worked out in order to diminish the detection limit in a pragmatic manner. Additionally, the application of bacteria detection by flow cytometry (BDFC) as a culture method is shown, i.e., the transfer of samples from PCs into a satellite bag, incubation of the latter at 37°C, and measuring the contaminating bacteria in a flow cytometer.