Application of IL-38 protein in preparation of medicine for treating ophthalmic diseases
An ophthalmic disease and protein technology, applied in the field of biomedicine, can solve the problems of blank and insufficient ophthalmic drugs, and achieve the effect of easy drug administration, easy and fast onset treatment plan, and low cost
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Embodiment 1
[0044] Preparation of IL-38 protein in different forms of embodiment 1
[0045] This example typically exemplifies the preparation of rhIL-38 protein and its fusion form of human serum albumin, and also exemplifies the method of expressing the target protein and the purification method of the Escherichia coli prokaryotic expression system and the Pichia pastoris expression system. The same or similar methods and steps can be used for other rhIL-38 protein fusion proteins, such as the form of rhIL-38 protein fusion IgG Fc fragment (rhIL-38-IgG Fc fusion protein), the form of fusion single domain antibody, human transferrin protein (transferrin) or other components that can extend the half-life-rhIL-38 protein fusion protein for preparation. The same or similar methods and steps can be used to express the target protein in other expression systems, such as mammalian cell expression system, insect expression system, etc.
[0046] (1) Preparation of rhIL-38 protein
[0047] The ...
Embodiment 2
[0055] Example 2: ELISA method to detect the binding of different forms of IL-38 protein to IL-36R
[0056] In this example, the binding ability of rhIL-38 protein, HSA-rhIL-38 fusion protein and rhIL-38-IgG Fc fusion protein to IL-36R protein was detected, and the EC50 was detected.
[0057] Experimental procedure
[0058] 1) Add 100 μL of carbonate coating solution containing 0.5 μg / mL IL-36R protein to the microwell plate, and coat overnight at 4°C;
[0059] 2) Tap off the liquid in the wells, add 300 μL of washing solution (0.1% PBST) to each well to wash the plate 3 times, soak the plate for 2 minutes each time and shake the plate gently to shake off the liquid in the plate, pat it on absorbent paper 3 times, and finally After one wash, pat dry on absorbent paper;
[0060] 3) Add 300 μL of blocking solution (PBS containing 1% BSA) to each well to seal the plate. Seal the plate with film and seal at 37°C for 1 hour;
[0061] 4) Repeat step 2);
[0062] 5) Add the stan...
Embodiment 3
[0074] Embodiment 3: ELISA method detects the combination of different forms of IL-38 protein and IL-1RI
[0075] In this example, the rhIL-38 protein, HSA-rhIL-38 fusion protein and rhIL-38-IgG Fc fusion protein were combined with IL-1RI protein, and their EC50 was detected.
[0076] Experimental procedure
[0077] 1) Add 100 μL of carbonate coating solution containing 10 μg / mL IL-1RI protein to the microwell plate, and coat overnight at 4°C;
[0078] 2) Tap off the liquid in the wells, add 300 μL of washing solution (0.1% PBST) to each well to wash the plate 3 times, soak the plate for 2 minutes each time and shake the plate gently to shake off the liquid in the plate, pat it on absorbent paper 3 times, and finally After one wash, pat dry on absorbent paper;
[0079] 3) Add 300 μL of blocking solution (PBS containing 1% BSA) to each well to seal the plate. Seal the plate with film and seal at 37°C for 1 hour;
[0080] 4) Repeat step 2).
[0081] 5) Add the standard curv...
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