Thanks for noticing.
Recently, I am going to do the semi-quantitative analysis for evaluating the inflammation situation in periodontal ligament tissue (PDL) of rat via immunostaining.
Now I have the antibody of the IL-1 beta and IL-6 , but I found the IL-1 beta and IL-6 are expressed in the cytoplasmic stained, not nuclear stained from the references
So, could I evaluate the inflammation by staining the IL-1 beta and IL-6?
If I want to stain the nuclear stained markers, do you have some suggestions about the marker?
And I want to do the Toluidine Blue staining for the Mast cell for the inflammation evaluation, is it feasible?
My samples are decalcified and embedded as paraffin blocks.
Looking forward to your reply.
Hallo Kai Li, different epitopes could be the reason for different expression. Compare the imunogens of your reference and your own antibody. You will find informations in the data sheet. In my opinion you can analyse the cytoplasmatic staining. You have to measure the intensity of the staining by densitometric measurements.
Good luck, Ute.
kai
I recommend to do different methods for assessment..
1. H&E histological assessment
of inflammation
Paraffinized sections (0.6 μm) from each patient (3–10 sections/patient) were stained with Gill’s hematoxylin V (MUTO, Tokyo, Japan) and 1% eosin alcohol solution (MUTO; H&E). The degree of inflammation was assessed according to a semi-quantitative scale. H&E-stained slides were examined under a light microscope (Olympus BX51; Olympus, Tokyo, Japan) in a double-blind manner by two histopathologists. Inflammation was scored as follows: mild inflammation,
2. Immunostaining assessment..
Slides were examined under an Olympus BX51 light microscope in a double-blind manner by two histopathologists. The staining intensity of the cytoplasmic staining should be scored as follows: 0, no staining; 1, mild staining; 2, moderate staining; 3, intense staining. In addition the percentage of positive cells within a 10 × 10 cm grid (area, 0.25 mm2; magnification, ×200) should be calculated ).
The percentage of positive cells should be counted regardless of staining intensity.
Statistical analysis
Statistical analysis have to performed using SPSS statistics software version 15.0 (SPSS, Inc., Chicago, IL, USA). The Morphological data can be presented as n (%) by inflammatory stage. The differences among the inflammatory stages can be compared using Fisher’s exact test. Spearman correlation analysis can be performed to identify correlations between the morphologic data and inflammatory stage. Results can be presented as the coefficient of correlation (r) and the corresponding P-value. Statistical assessments will be two-tailed and significance is set at P
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