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Inertial microfluidics combined with selective cell lysis for high throughput separation of nucleated cells from whole blood
KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.ORCID-id: 0000-0001-5199-0663
KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.
KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi.ORCID-id: 0000-0002-0242-358X
2017 (Engelska)Ingår i: RSC Advances, ISSN 2046-2069, E-ISSN 2046-2069, nr 47, s. 29505-29514Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

The ability to rapidly analyze and extract information from peripheral blood cells has the potential of providing a wealth of new information about immune function and general health of the patient. In spite of the tremendous progress achieved in the field of leukocyte analysis, one of the major impediments for routine analysis is the enrichment of cell populations from heterogeneous sources such as blood, as the currently used techniques tend to be laborious. Moreover, the isolation of small and transient cell populations in blood, like circulating tumor cells during cancer metastasis, is even more challenging. Here, we report an integrated device for label-free continuous flow separation of nucleated cells from unprocessed whole blood at high throughput. The method utilizes exposure to hypotonic buffer to completely remove red blood cells and at the same time a size increase of nucleated cells for inertial focusing and separation in spiral microchannel. Using an integrated device with two outlets, we isolated total leukocytes at a high yield of 99%. Furthermore cancer cells spiked into whole blood could be separated at a yield of 88% while 80% of leukocyte could be depleted into separate outlet by simply changing the resistance between the two outlets. Finally, using a three-outlet integrated device, we demonstrate fractionation of leukocyte into subpopulation. The device continuously separates granulocytes at a purity of 86%, monocyte at a purity of 43% and lymphocytes at a purity of 91% simultaneously. Finally, a cell activation study of the immune system using blood from healthy subjects, stimulated ex vivo with lipopolysaccharides (LPS), confirmed that the high operational flow rate of the device does not alter the activation levels of leukocytes or introduce artifacts. Hence, the simple, high-throughput and low-cost integrated device requiring neither external force fields nor mechanical parts to operate should readily be applicable to sort nucleated cells as stand-alone and/or as integrated lab-on-a-chip devices with high-throughput requirements.

Ort, förlag, år, upplaga, sidor
RSC Publishing, 2017. nr 47, s. 29505-29514
Nyckelord [en]
inertial microfluidics, nucleated cells, selective cell lysis
Nationell ämneskategori
Medicinteknik
Forskningsämne
Bioteknologi
Identifikatorer
URN: urn:nbn:se:kth:diva-203241DOI: 10.1039/C7RA02992FScopus ID: 2-s2.0-85021694374OAI: oai:DiVA.org:kth-203241DiVA, id: diva2:1081726
Forskningsfinansiär
BarncancerfondenEU, FP7, Sjunde ramprogrammetVetenskapsrådet
Anmärkning

QC 20170320

Tillgänglig från: 2017-03-14 Skapad: 2017-03-14 Senast uppdaterad: 2018-09-19Bibliografiskt granskad
Ingår i avhandling
1. Microfluidic based isolation of circulating tumor cells from whole blood for cancer diagnostics
Öppna denna publikation i ny flik eller fönster >>Microfluidic based isolation of circulating tumor cells from whole blood for cancer diagnostics
2017 (Engelska)Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
Abstract [en]

Detection of circulating tumor cells (CTC) in peripheral blood is indicative of early recognition of tumor progression and such an important biomarker for early diagnosis, staging, monitoring and prognosis of cancer. However, CTC are found in very low concentrations and reliable isolation of these rare cells is challenging. Microfluidics enables precise manipulation of fluids and cells and is ideal for cell sorting methods for clinical diagnostics. The thesis contributes towards the development of microfluidic based CTC isolation methods from peripheral blood. The methods are based on size and immunoaffinity. The first part of the thesis describes the phenomenon of inertial focusing for size based cell separation at high throughputs. In paper 1, we demonstrate continuous filtration of leukocytes from diluted blood, with an efficiency of 78% at a flow rate of 2.2ml/min. In the paper 2, separation of total and subpopulation of leukocytes with a purity of 86% for granulocytes and 91% for lymphocytes is demonstrated. Furthermore, cancer cells spiked into whole blood could be separated at a yield of 88%. Finally, in paper 3 and 4 we unravel parts of the unexplored elasto-inertial microfluidics and was utilized to precisely focus the cells, as part of an integrated optofluidic micro flow cytometer device, capable to simultaneously measure fluorescence and scattering of cells and particles at a rate of 2500 particles/sec (paper 4). Second part of the thesis focuses on acoustophoresis. In (paper 5), a multifunctional acoustic microdevice was developed for isolation of cancer cells from red blood cells with a separation efficiency of 92.4% and trapping efficiency of 93%. In (paper 6), microbubbles activated acoustic cell sorter was developed for affinity based cell separation. As a proof of principle, cancer cells in a suspension were separated at an efficiency of 75%. In the third part, using cellulose nano fibrils (paper 7), we demonstrate efficiently capture and release of cancer cells at a release efficiency of 95%. Finally, a novel, single step self-assembly of spider silk proteins is introduced inside microfluidic channels for effective capture of cancer cells with 85% capture efficiency and subsequent release of captured cells with 95% release efficiency (paper 8). The novel recombinant silk modified microfluidic device was validated using pancreatic cancer patients. In summary, we have developed different microfluidic based isolation technologies for the capture and characterization of CTC.

Ort, förlag, år, upplaga, sidor
stockholm: KTH Royal Institute of Technology, 2017. s. 109
Serie
TRITA-BIO-Report, ISSN 1654-2312 ; 2017:7
Nationell ämneskategori
Medicinteknik Medicinsk bioteknologi
Forskningsämne
Bioteknologi
Identifikatorer
urn:nbn:se:kth:diva-203889 (URN)978-91-7729-311-8 (ISBN)
Disputation
2017-04-13, Gardaulan, Folkhälsomyndigheten, Nobels väg 18, Solna, Stockholm, 10:00 (Engelska)
Opponent
Handledare
Forskningsfinansiär
EU, FP7, Sjunde ramprogrammetBarncancerfondenVetenskapsrådet
Anmärkning

QC 20170321

Tillgänglig från: 2017-03-20 Skapad: 2017-03-20 Senast uppdaterad: 2017-03-23Bibliografiskt granskad

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Ramachandraiah, Harisha

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Ramachandraiah, HarishaSvahn Andersson, HeleneRussom, Aman
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