Comparison of CD45+ Depletion Methods for Enrichment of Disseminated Tumor Cells in Bone Marrow Samples
Poster Dec 07, 2017
Birte Möhlendick, Imke Hoffmann, Swetlana Seidschner, Sarah Schumacher, Alexander Smyczek, Carina Vaerst, Wolfram Trudo Knoefel, Nikolas Hendrik Stoecklein
Disseminated tumor cells (DTC) in bone marrow (BM) samples are, with a frequency of only one DTC per 1x106 cells, quite rare. DTCs are usually detected utilizing immunofluorescence staining of 1-2x106 mononuclear cells (MNC) and consecutive microscopic analysis. To increase the detection rate of DTCs a higher number of cells should be analyzed. Since microscopic detection of these cells is very laborious DTC enrichment strategies are needed. Here, we investigated the efficiency of three different depletion strategies of CD45+ cells for DTC enrichment in BM samples.
CD45+ cell depletion was performed on BM samples from cancer patients with different tumor entities. Immunomagnetic depletion (DYN) of CD45+ cells was performed after Ficoll density gradient centrifugation on 1x107 MNCs utilizing CD45 Dynabeads® (Invitrogen) with i) 4x107 beads/ml cell suspension (DYNLO) and ii) 1x108 beads/ml cells suspension (DYNHI). Antibody-based depletion (ROS) of CD45+/CD66b+ cells was performed on 1x108 BM cells with the RosetteSep® Human CD45 Depletion Cocktail (Stemcell Technologies). For the different strategies at least 60 samples were analyzed, the depletion efficiency was calculated and DTC numbers were extrapolated to compare the respective methods.
We were able to detect DTCs in 32.7% (DYNLO), 12.5% (DYNHI) and 66% (ROS) of the patient samples, respectively. CD45+ cells could be depleted most efficiently up to 440-fold using ROS, followed by 44-fold with DYNHI and 20-fold using DYNLO. DTC numbers were significantly higher (p<0.0001) in the ROS samples (mean=3.69) with up to 21 DTCs per 1x106 cells. With the immunomagnetic procedures DYNLO samples (mean=0.48) had slightly higher DTC counts (8.33 vs 6.35 DTCs per 1x106 cells) compared to DYNHI (mean=0.39).
In our hand the ROS procedure showed the best results for DTC enrichment from BM samples. This method enables the analysis of a high number of BM cells since it depletes unwanted CD45+ cells most efficiently and thus increases DTC detection rates in the patient samples.
Treatment Options for Chronic Parvovirus Viremia in Pediatric Heart Transplant Patients in a Tertiary Care CenterPoster
This abstract discusses three cases of pediatric heart transplant patients who suffered from parvovirus (B19) infection. Of these patients, two ( B & C) responded well to standard intravenous Ig therapy. Patient A however, did not respond to standard treatment and was begun on subcutaneous Ig, which effectively diminished his viral load. Thus, subcutaneous Ig infusions might serve as a second line treatment for transplant patients with parvovirus who do not respond well to the standard approach.READ MORE
T-helper cell phenotype expression in cutaneous lesions of angioimmunoblastic T-cell lymphomaPoster
Angioimmunoblastic T-cell lymphoma (AITL) is a common type of peripheral T-cell lymphoma. AITL can be missed until lymphadenopathy develops in patients initially presenting with skin lesions, as skin biopsy may lack conclusive findings. Our case highlights the extranodal presentation of AITL with cutaneous lesions displaying the TFH phenotype.READ MORE
The Utility of Pediatric Asthma Severity Scores in Disposition Decisions for Status AsthmaticusPoster
We aimed to determine if the Pediatric Asthma Severity Score (PASS) predicts a patient's appropriate level of care in the inpatient setting, with special attention to patients who required rapid transfer to another level of care after initial disposition and/or hospital readmission.READ MORE
Comments | 0 ADD COMMENT
2nd Annual Artificial Intelligence in Drug Development Congress
Sep 20 - Sep 21, 2018