Leukocyte adhesive function assays, devices and/or uses
A white blood cell and cell technology, applied in the field of measurement, can solve the problems of lack of ability to quantitatively evaluate the adhesion function of specific adhesion molecules and limited applications
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example 1
[0297] Example 1: Mn 2+ Activation of α4β1 integrin adhesion function.
[0298] In this exemplary embodiment, in order to evaluate the ability of leukocytes to interact with endothelial VCAM-1, a microfluidic system consisting of a microfluidic pump and a microfluidic chip was used to simulate in vitro blood microcirculation. The bottom of the chip was pre-coated with VCAM-1 protein (10 μg / ml) overnight at 4°C, and then whole blood was perfused through the channel at a flow rate of 10 μl / min driven by a microfluidic pump. The protocols used are set in Appendix I unless otherwise stated.
[0299] Simultaneous detection of VCAM-1-dependent recruitment of CD4, CD8, and CD15 cells
[0300] In this example, to avoid cell separation and achieve simultaneous detection of multiple specific leukocyte subsets, fluorescently labeled antibodies against specific leukocyte membrane markers were added to human whole blood prior to performing the flow assay. Such as figure 1 As shown in A, ...
example 2
[0305] Example 2: Using Multiple Membrane Markers to Identify Specific Leukocyte Subgroups
[0306] According to certain exemplary embodiments, the present embodiments relate to identifying CD15 and CD16 double positive neutrophils in an assay. Experiments were performed following the protocol set forth in Appendix I with the following modifications:
[0307] 1. Add 3 μl of anti-human CD15-APC (BD, catalog number: 551376) and 2 μl of anti-human CD16-BV510 (BD, catalog number: 563830) into 100 μl of human whole blood before using in the assay, and Incubate for approximately 5 minutes at room temperature,
[0308] 2. Use a macro called "Multi_Channel.ijm" in Fiji software to track CD15CD16 double positive cells.
[0309] Most interacting CD15 cells are neutrophils. Eosinophils are a small population of granulocytes known not only to have high expression levels of α4 integrin but also to be positive for CD15 expression. To determine whether the interacting CD15 cells are neut...
example 3
[0311] Example 3: Semi-quantitative assessment of the basal inflammatory state of α4β1 integrins
[0312] This example is directed to illustrating semi-quantitative assessment tools that may be used in certain exemplary embodiments. In disease subjects, increasing the fraction of activated α4β1 integrin molecules compared to healthy subjects results in enhanced leukocyte binding to α4β1 integrin endothelial ligands (e.g., VCAM-1), and leukocyte recruitment and / or increased inflammatory response.
[0313] The following criteria are used to define one or more healthy subjects:
[0314] 1. Apparently healthy as determined by medical evaluation including medical history.
[0315] 2. Women who are not pregnant or are currently breastfeeding
[0316] 3. Not diagnosed with any autoimmune, inflammatory, blood and vascular conditions
[0317] 4. Not currently taking prescription drugs, except birth control pills
[0318] 5. Currently not taking over-the-counter drugs that may af...
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