Analyzing Macrophages
Pariza F -
This week I was working with two sets of data. One was the brain slides with microglia staining, which I have been working on for the past couple of weeks. The other set of images I am working on are DAPI stainings, SOX9 stainings, S100B stainings and GFAP stainings. DAPI stainings are mainly used for identifying nuclear DNA, while GFAP, SOX9, and S100B stainings are used to identify astrocytes, astrocyte development, and mature astrocytes in the brain.
The purpose of staining nuclear DNA is to see the number of live cells that remain active after the brain experiences a traumatic injury. Seeing less nuclear DNA stains on a brain slide tissue is a major sign of neurodegeneration. We used this in our lab to see the extent of a mild traumatic brain injury in a rat.
GFAP, SOX9, and S100B stainings were done through immunoflorescence. This staining technique allowed us to have all three markers on one brain slide in different colors. The markers showed us all the things related to astrocytes in these brains. Now, you are probably wondering what astrocytes do… Astrocytes maintaining brain homeostasis and protect the brain from injury by regulating the blood-brain barrier. The blood-brain barrier protects the brain from the body by not letting the symptoms of the body affect the brain. For example, if the flu affected your brain it would cause severe damage in your health.
Besides that, I wanted to go further explain the immunoflorescence staining protocol. Immunofluorescence staining is a technique used to visualize specific proteins or antigens in cells or tissues using fluorescently labeled antibodies. It relies on the principle of antigen-antibody binding and fluorescence microscopy. There are two main types: direct and indirect immunofluorescence. In the direct method, a primary antibody conjugated with a fluorescent dye binds directly to the target antigen. In the indirect method, an unlabeled primary antibody binds to the antigen, followed by a fluorescently labeled secondary antibody that recognizes the primary antibody, amplifying the signal.
Below are some of the images:
This one is the DAPI staining:
This one is the GFAP, SOX9, and S100B staining, and each color is magenta, green, and red, respectively: