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Specific marker Glut3 (Glucose Transporter 3) of embryonic stem cells and application thereof

An embryonic stem cell, specific technology, applied in the field of cell biology and immunology, can solve the problem of the limited number of surface markers on the embryonic stem cell membrane, and achieve the effect of high specificity and sensitivity

Inactive Publication Date: 2013-07-17
杭州华安生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Moreover, the specificity of these markers has certain limitations: they are also expressed in some adult stem cells, normal tissues or cancer cells, and the number of membrane surface markers specific to embryonic stem cells is even more limited

Method used

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  • Specific marker Glut3 (Glucose Transporter 3) of embryonic stem cells and application thereof
  • Specific marker Glut3 (Glucose Transporter 3) of embryonic stem cells and application thereof
  • Specific marker Glut3 (Glucose Transporter 3) of embryonic stem cells and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Example 1 Preparation of Glut3 rabbit monoclonal antibody

[0020] Using mouse embryonic (mES) stem cells as the immunogen, 1×10 8 Three-month-old New Zealand white rabbits were immunized with mES, rabbit spleen cells were isolated, and myeloma cells 240E-W2 (purchased from Epitomics) were fused to screen out the hybridoma cell line ZJUESRMAB39 (preservation number CGMCC NO.) that produces Glut3 rabbit monoclonal antibodies. 7301), applied to the identification and separation of stem cells.

Embodiment 2

[0021] Example 2 Application of Glut3 as a marker for embryonic stem cells

[0022] 1.1 Glut3 rabbit monoclonal antibody recognizes natural conformational epitopes and can be used for flow cytometric screening of embryonic stem cells

[0023] The statistical results of flow cytometry data show that the expression of Glu3 decreases significantly with ES cell differentiation.

[0024] Such as figure 1 It shows that 70% of intact mES can be positively stained by the Glut3 rabbit monoclonal antibody prepared in the example, while only 3% of mES cells induced to differentiate by 10 μmol / L at RA (retinoic acid) are positively stained.

[0025] Flow cytometry (FC): undifferentiated mES cells without feeder layer and differentiated by 10μmol / L at RA were separated into single cells; incubated with Glut3 rabbit monoclonal antibody and negative control antibody respectively; FITC-goat antibody was added Incubate with rabbit secondary antibody; wash with PBS; observe by flow cytometry.

[0026] I...

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Abstract

The invention provides a novel specific marker Glut3 (Glucose Transporter 3) of embryonic stem cells and application thereof. A Glut3 polyclonal antibody or a rabbit monoclonal antibody Glut3 prepared by directly taking the embryonic stem cells as immunogen is used as stem cell identification reagents; the specificity and the sensitivity of detection are higher; furthermore, the specific marker Glut3 can be used for separation and activity detection of the embryonic stem cells. An embryonic stem cell detection kit containing the Glut3 polyclonal antibody or the rabbit monoclonal antibody Glut3 can be used for identifying the totipotency of the embryonic stem cells and also can be used for detecting activity of stem cells simultaneously; therefore, functions of the detection kit are expanded.

Description

Technical field [0001] The invention belongs to the field of cell biology and immunology, and specifically relates to embryonic stem cell specific marker Glut3 and its application. Background technique [0002] Embryonic stem cells (ES cells) refer to normal diploid cells that are separated from the inner cell mass of the embryo and can be cultured in vitro for a long time and maintain the ability of self-renewal and differentiation into all three germ layer cells. The high self-renewal ability and differentiation totipotency of embryonic stem cells are important characteristics that distinguish it from other cells. Its specific molecular markers are of great significance for the identification and isolation of embryonic stem cells and the study of self-renewal and differentiation mechanisms. Commonly used embryonic stem cell-specific markers include transcription factors OCT4, NANOG, REX1, etc.; surface markers SSEA-1, SSEA-3, TRA-1-60, CD9, etc.; these embryonic stem cell marke...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/47C12N5/20G01N33/68G01N33/577C12R1/91
Inventor 张铭侯倩女宋秀丽顾斌张佳蓉谭舟
Owner 杭州华安生物技术有限公司
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