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. Author manuscript; available in PMC: 2017 Dec 1.
Published in final edited form as: Cytometry A. 2016 Nov 22;89(12):1073–1083. doi: 10.1002/cyto.a.23019

Table 3.

DLD microchip processing of immunostained unlysed blood using optimized buffer resulted in high recovery of washed WBCs with almost 10,000-fold depletion of RBCs.

Experimenta Fraction
Analyzed
Volume (μL)b RBC Count (× 106 Cells/μL) %RBC
Depleted
%RBC Depleted Flow Cytometryc WBC Count (× 103 Cells/μL)d Total % WBC Recoverye % Granulocytesf % Monocytesg % Lymphocytesh % B-Cellsi % T-Cellsj Run Time (Mins)
With Optimized Buffer
1 Input 200 2.66 4.0 87 +/− 5% 54.0 5.3 40.7 9.3 75.5 21.0
Product 289 0.01 99.62% 99.993% 2.4 52.6 14.5 32.9 11.9 77.1
2 Input 200 2.01 3.2 88 +/− 6% 47.6 9.8 42.8 9.0 85.9 16.0
Product 255 0.00 100.00% 99.974% 2.2 52.2 10.4 38.2 10.3 87.3
3 Input 200 2.45 2.6 78 +/− 7% 60.3 8.6 30.0 15.7 67.3 17.0
Product 240 0.00 100.00% 99.997% 1.7 64.4 7.4 29.1 17.0 63.8
4 Input 200 2.23 4.7 96 +/− 4% 60.3 9.3 30.3 13.0 79.3 18.0
Product 281 0.01 99.55% 99.993% 3.2 61.8 8.2 28.4 14.6 74.9
5 Input 200 2.06 2.0 81 +/− 9% 42.3 7.9 46.9 9.2 77.0 18.0
Product 248 0.00 100.00% 99.961% 1.3 40.0 10.5 47.4 8.8 77.8
6 Input 200 2.19 2.1 98 +/− 8% 53.6 7.3 38.4 9.4 64.3 20.0
Product 241 0.00 100.00% 99.991% 1.7 51.2 8.0 39.8 9.0 65.6
Mean Depletion or Recovery or Run Time 99.8 + /− 0.09% 99.985+/− 0.006% 88 +/− 3% 18.3+/-0.8
Mean Difference {Product vs. Input) 0.7+/− 1.3% 1.8 +/− 1.6% −2.2+/− 1.4% 1.0 +/− 0.5% −0.5 +/− 1.1%
Control with BSA Buffer
1 Input 200 2.68 3.8 77 +/− 5% 54.0 5.3 40.7 9.3 75.5 22.0
Product 278 0.01 99.63% 99.991% 2.1 53.0 11.9 35.1 5.5 82.0
a

“Control with BSA Buffer” Experiment 1 used BSA buffer and the same donor blood sample, and was run at the same time as Experiment 1 “with Optimized Buffer”. Experiments 1 – 6 with “Optimized Buffer” used the buffer containing Polyoxamer instead of BSA.

b

As in Tables 1 and 2.

c

%RBC depletion was determined using relative ratios of RBC/WBC for both the Coulter Counter and flow cytometry approaches.

d,e

As in Tables 1 and 2.

f

Granulocytes were identified by flow cytometry as DRAQ5+ signals with characteristic high FSC and SSC.

g

Monocytes were identified as DRAQ5+ with characteristic intermediate FSC and SSC.

h

Lymphocytes were identified as DRAQ5+ with characteristic low FSC and SSC.

i

B cells were identified as DRAQ5+/CD19+ with FSC and SSC of lymphocytes.

j

T cells were identified as DRAQ5+/CD3+ with FSC and SSC of lymphocytes.