(20,) (21) The expression of the surface marker of the stem cells was analyzed via the incubation of at least 100 000 cells with fluorescence-labeled monoclonal antibodies against CD29
, CD34, CD44, CD45, and CD90 (Sigma, China).
The gene of Oct-4, Rex-1, CD29
, CD44, CD71, and CD73 were identified by RT-PCR and immunofluorescence technique.
Results: ADSCs were successfully isolated from rat adipose tissue as identified by expression of typical surface markers CD29
, CD44, CD90, and CD45 in flow cytometry.
MenSCs and BMSCs expressed MSCs markers including CD9, CD10, CD29
, CD44, CD73 and CD105, they were also negative for hematopoietic markers, CD34, CD38, CD45 and CD133.
Subsequently, all MSCs at passage 4 homogeneously showed positive expression for mesenchymal markers CD29
(ADSC, 99.53% [+ or -] 0.12%; UCMSC, 98.7%0 [+ or -] 0.30%; EMSC, 90.75% [+ or -] 0.05%), CD90 (ADSC, 97.60% [+ or -] 1.21%; UCMSC, 98.4% [+ or -] 0.33%; EMSC, 92.55% [+ or -] 0.05%), CD73 (ADSC, 98.75% [+ or -] 0.85%; UCMSC, 97.9% [+ or -] 0.5%; EMSC, 99.8% [+ or -] 0.2%), and CD105 (ADSC, 98.95% [+ or -] 0.45%; UCMSC, 93.4% [+ or -] 1.02%; EMSC, 96.95% [+ or -] 0.45%) and were negative for CD31 (<1%), CD34 (<2%), CD45 (<2%), CD14 (<1%), CD19 (<1%), and HLA-DR (<1%)
The activated T cells in the airway wall are associated with inflammation of asthma [45, 46], and the subsets of T cell antigens have attracted extensive attention by researchers, such as the T cells of CD4 +(T helper) [47-51], CD8+ [52-54], CD25+ , CD28 [56, 57], CD29
[58, 59], CD39+, and CD73 [60-63].
The immunophenotypes of these cells were persistently positive for CD29
, CD44, and CD90 but negative for CD34, CD45, and CD11b for more than three passages.
The cells were incubated with antimouse/rat CD29
antibody (Biolegend, USA), phycoerythrin/CD44 antibody (Abcam, United Kingdom), PE-Cy[TM]5 mouse anti-rat CD45 antibody (BD Biosciences, USA), antirat/mouse CD90.1 (Thy-1.1) antibody (E-Bioscience, USA), and anti-mouse endoglin/CD 105 antibody (R&D Systems, USA) for 20 min.
Third-passage DFSCs were incubated with the antibodies of positive markers for MSCs of human CD29
allophycocyanin (APC), CD73 phycoerythrin (PE), CD105 PE, and CD146 fluorescein isothiocyanate (FITC) and with the antibodies of negative markers of CD14 PE, CD34 FITC, CD45 APC, and HLA-DR PE (BD Biosciences, USA) for 15 min at room temperature in the dark.
The cells presented high positivity for the following expressions: CD29
(98.2 [+ or -] 1.1%), CD44 (76.65 [+ or -] 2.55%), CD73 (83.2 [+ or -] 15%), and CD90 (90.05 [+ or -] 8.65%).
Immunophenotype was analyzed using a flow cytometer with the following fluorescein isothiocyanate (FITC), allophycocyanin (APC), or phycoerythrin (PE) antibodies: CD29
, CD44, CD90 (eBioscience, USA), CD34 (Abcam, Cambridge, MA, USA), and CD45 (BD Pharmingen, USA).