presented by: casey hua, danyi wu
DESCRIPTION
Isolating highly enriched populations of circulating epithelial cells and other rare cells from blood using a magnetic sweeper device. - PowerPoint PPT PresentationTRANSCRIPT
Isolating highly enriched populations of circulating
epithelial cells and other rare cells from blood using a magnetic
sweeper deviceAmirAli H. Talasaz, Ashley A. Powell, David E. Huber,
James G. Berbee, Kyung-Ho Roh, Wong Yu, Wenzhong Xiao, Mark M. Davis, R. Fabian Pease, Michael N.
Mindrinos, Stefanie S. Jeffrey, and Ronald W. Davis
Presented by: Casey Hua, Danyi Wu
Motivation
• Molecular Profiling Applications
• Improvement of:• Cancer prognosis/treatment rare circulating
epithelial cells (CEpCs)• Immune monitoring antigen-specific
lymphocytes• Prenatal diagnosis fetal cells
Motivation
Problem: Current CEpC isolation platforms lack purification efficiency
Solution: MagSweeper–High capture rate–High purity–Enriches target cells (108x)
MagSweeper Process1. Pre-label diluted blood samples with magnetic particles
2. 3. 4.
5. Repeat to eliminate background cells improves purity at some cost to capture efficiency
MagSweeper Simulation
Magnetic flux density
Magnetic force-driven cell velocity
field
Cell capture boundary
MagSweeper Simulation cont’dFluid
velocity/motionCell trapping profiles
Gap of uncaptured particles due to fluid motion pushing particles
ahead of magnet
• Enrichment Studies1. Rare Immunological Model Cells from a Mixed Sample (HLA-
A2)• Capture Efficiency • Purity
2. MCF7 Cancer Cells Spiked in Blood Samples• Performance: isolating Circulating Epithelial Cells (CEpCs)
3. CEpCs from Breast Cancer Patients• Application: CEpC isolation from Epithelial Cancers patients
• Perturbation of Cellular Gene Expression
MagSweeper Performance
(1) Rare HLA-A2+ cells in Mixed Samples
Enrichment by ~2.5x105
(2) MCF7 Cancer Cells Spiked in Blood Samples
MCF7 cells isolated by MagSweeper with brightfield and fluorescence imaging.
Combining MagSweeper enrichment with visual inspection and extraction of cells based on size:• increase cell purification of the MCF7 cells from 51% to 100% purity• similar cell capture rates from 62%± 7% of 59%± 27%
(3) CEpCs in Breast Cancer Patients
• Found CEpCs in 47 of 47 samples from patients with metastatic disease
• 12±23 CEpCs per 9mL blood sample
MagSweeper isolation protocol does not perturb gene expression
Fig. S2. Fold changes in logarithmic scale between the gene expression levels of cultured cells before and after MagSweeper protocol.
minimal changes in cell profiles: 94% of probe sets have changes <50%
Advantages of MagSweeper• High enrichment• No prior processing of blood required• Flexibility in starting sample volume & process
scalability• Further Optimization:
– Extension of the sweep to capture remaining particles in gap left by fluid motion
– Refine magnet shape to yield more uniform magnetic field gradient
Significance• Diagnose malignancy from blood
– Improved cancer prognosis: CEpC numbers correlate better with cancer progression than imaging, lymph node biopsy, and serum biomarker analysis
• Enrich CEpCs for molecular analysis– Identify surrogate endpoints for evaluation in clinical trials
• Extract individual CEpCs• characterization of single cells for subpopulation studies on
heterogenous CEpCs