Neutrophil Extraction via Density Gradient Centrifugation
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Solution Overview
Problem
Current methods for detecting neutrophil chemotaxis are time-consuming and inefficient, making it difficult to accurately diagnose and treat immune-related diseases.
Innovation Solution
A method involving blood sample processing using EDTA anticoagulant, centrifugation, density gradient centrifugation, and erythrocyte lysis to quickly extract and analyze neutrophil chemotactic function, which includes steps like adding glucose, centrifuging, re-suspending with ion-free Hank's solution, and lysing erythrocytes to isolate neutrophils for chemotactic assessment.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If conventional neutrophil extraction methods are used, then neutrophils can be obtained for detection, but the extraction process is time-consuming and inefficient
Solution Approach 1:
The patent applies preliminary action by adding glucose to the blood sample before centrifugation to induce neutrophil aggregation and separation from other blood components. This pre-treatment step accelerates the extraction process by naturally concentrating neutrophils in a specific layer, eliminating the need for multiple sequential separation steps and reducing overall detection time while maintaining extraction efficiency
2Ease of manufacture
If conventional extraction methods are used, then neutrophils can be isolated, but the process requires multiple complex steps including centrifugation and lysis
Solution Approach 1:
The patent merges multiple extraction steps into a single centrifugation process. By combining the separation of neutrophils from plasma and the aggregation of neutrophils into one centrifugation step following glucose addition, the method simplifies the procedure from multiple sequential operations to a single integrated process, reducing procedural complexity while maintaining extraction effectiveness
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This method significantly reduces the time required for neutrophil extraction and detection, improving extraction speed and accuracy, allowing for faster and more efficient analysis of neutrophil chemotaxis.
Implementation Method 1
centrifuging the supernatant at a centrifugal force of 400 g and 20° C. for 10 minutes
Implementation Method 2
slowly adding a polysucrose solution from the bottom of the centrifuge tube to perform density gradient centrifuging treatment
Implementation Method 3
lysing erythrocytes in the cell aggregates by using sterile water
Data Source
AI summary
Disclosed is a method for detecting chemotaxis of neutrophil, including: mixing a blood sample and an equal volume of glucose, mixing, and standing; taking a supernatant of the blood sample after standing into a centrifuge tube, and centrifuging; adding 1× calcium and magnesium ion-free HBSS into a centrifuged blood sample to re-suspend cell aggregates at bottom; uniformly blowing and dispersing a re-suspended bottom cell aggregates, and slowly adding a polysucrose solution from the bottom of the centrifuge tube to perform density gradient centrifuging treatment; dividing a density gradient centrifuged solution into three layers, pipetting a clear liquid at an upper layer and a PBMC layer at a middle layer to obtain cell aggregates at a bottom layer; lysing erythrocytes in the cell aggregates, then successively adding 2× calcium and magnesium ion-free HBSS and 1× calcium and magnesium ion-free HBSS, mixing, and centrifuging again to obtain neutrophils.