Olfactory receptor AI1 antibody

Antibody preparation process:

1. Materials and reagents

a. Extracted animal Ig

b. Freund's adjuvant and Freund's adjuvant

d. Experimental animal rabbit

e. Other materials and reagents

2. Select the experimental live organism.

3. Conduct animal immunization experiments.

4. Take a blood sample for testing to see the immune effect.

5. If the immunization is successful, kill the experimental live organism and collect all serum.

6. Purify the antibody.

7. Identify antibodies. Fetal bovine serum (sterile collection)
Detailed introduction:

English name:Hour1

English alias:Calciumreleaseactivatedcalciumchannelprotein1; Calciumreleaseactivatedcalciummodulator1; Calciumrelease-activatedcalciumchannelprotein1; CRACM1; CRACM1; CRCM1; CRCM1_HUMAN; FLJ14466; Orai1; ORAIcalciumreleaseactivatedcalciummodulator1; orai1; ORAT1; Proteinorai1; Proteinorai-1; TMEM142A; TMEM142A; Transmembraneprotein142A.

cross-reaction:Human, Mouse, Rat, Dog, Pig, Cow, Horse, Rabbit, Sheep

mark:Unconjugated

Antibody source:Rabbit

Antibody type:Polyclonal

Immunogen:KLHconjugatedsyntheticpeptidederivedfromhumanOrai1

Protein cell localization: Cell membrane

Purification method:affinitypurifiedbyProteinA

subtype:IgG

trait:Liquid

concentration:1mg/ml

Storage liquid:0.01MTBS(pH7.4)with1%BSA,0.03%Proclin300and50%Glycerol.

Storage conditions:Shippedat4℃. Storeat-20°Cforoneyear. Avoidrepeatedfreeze/thawcycles.
The company's products are only used for scientific research
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operating steps:

1. Dewaxing and hydration. Soak the paraffin sections in fresh xylene for 15 minutes and repeat three times. After removing excess liquid, immerse in anhydrous ethanol, 95% ethanol, 90% ethanol, 80% ethanol, and 70% ethanol for 5 minutes each, and rinse with distilled water (or tap water) for 5 minutes. Wash the slices with PBS buffer (reagent 1, dissolve the dry powder in 1L of deionized water) for 5 minutes and repeat three times.

2. Antigen repair. Place the dewaxed and hydrated tissue slices on a high-temperature resistant plastic slice rack in a beaker (or repair kit), add an appropriate amount of repair solution 1 x sodium citrate antigen repair solution (reagent 2, dilute 100 x sodium citrate antigen repair solution with deionized water to 1 x sodium citrate antigen repair solution), and the liquid level should be immersed in the sliced tissue to a certain height. Boil the repair kit in boiling water for 15 minutes, then remove the slide and let it cool naturally to room temperature. Wash the slices with PBS buffer for 5 minutes and repeat three times.

AttentionWhen antigen repair is performed, the repair solution must ensure that the slices are always immersed in the liquid. Generally, the amount of repair solution is about 800mL per rack.
3. Block endogenous peroxidase. Wipe the glass slide dry with absorbent paper, draw circles around the tissue with an immunohistochemical pen, and drop 50 μ L of endogenous peroxidase inhibitor (reagent 3) onto the sliced tissue to block endogenous peroxidase. Incubate at room temperature for 30 minutes, rinse the slices with PBS buffer for 5 minutes, and repeat three times.

4. Serum blockade. Wipe the glass slide dry with absorbent paper, add 50 μ L of normal goat serum working solution (reagent 5) dropwise, and seal at 37 ℃ for 20 minutes to reduce non conformities

Specific staining.

5. Incubation of primary antibodies. Dilute the primary antibody solution into a working solution using antibody diluent (reagent 4), dry the liquid around the glass slide tissue with absorbent paper, add an appropriate amount of antibody working solution dropwise, and incubate overnight at 4 ℃ or at 37 ℃ for 1-2 hours in a wet box.

6. Reheat. After overnight incubation at 4 ℃, take out the sample from the refrigerator and incubate it at room temperature for 15 minutes for reheating (this step is selected for antibody incubation at 4 ℃ overnight, if incubated at room temperature, proceed directly to the next step of cleaning). Wash the slices with PBS buffer for 5 minutes and repeat three times.

7. Secondary antibody incubation. After drying the slices with absorbent paper, add 50 μ L of labeled sheep anti rabbit IgG (reagent 6) dropwise, incubate at 37 ℃ for 20 minutes, and use PBS

Wash the slices with buffer solution5 minutes, repeat three times.

8. Triple antibody incubation. After drying the slices with absorbent paper, add 50 μ L of horseradish enzyme labeled streptavidin and incubate (reagent 7). Incubate at 37 ℃ for 20 minutes, rinse the slices with PBS buffer for 5 minutes, and repeat three times.

9. Color rendering. Shake off PBS buffer and dry the slices with absorbent paper; Add 50 μ L of freshly prepared DAB working solution (reagent 8: reagent 9: reagent 1=1:1:18 ratio) to each slice, incubate for 3-5 minutes, and observe the staining results under an optical microscope. Do not darken the color; After color development, rinse the slices with tap water to terminate color development.

10. Re dyeing. Add an appropriate amount of hematoxylin staining solution (reagent 10) dropwise for counterstaining, incubate for 1-5 minutes, rinse with tap water for 5 minutes, and differentiate by adding hydrochloric acid and alcohol dropwise

Liquid differentiation30 seconds, rinse with tap water.

11. Dehydration sealing. Soak the glass slides in 70% ethanol, 80% ethanol, 90% ethanol, 95% ethanol, and anhydrous ethanol for 5 minutes each, and then soak the slides in xylene for 15 minutes, repeating this process three times. Take out the glass slide and let it dry, then seal it with neutral gum.

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