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ASL antibody - middle region (ARP41667_T100)

100 ul
$229.00
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Conjugation Options

ARP41667_T100-FITC Conjugated

ARP41667_T100-HRP Conjugated

ARP41667_T100-Biotin Conjugated

Gene Symbol:
ASL
NCBI Gene Id:
435
Official Gene Full Name:
Argininosuccinate lyase
Protein Name:
Argininosuccinate lyase
Swissprot Id:
P04424
Protein Accession #:
NP_001020115
Nucleotide Accession #:
NM_001024944
Alias Symbols:
ASAL
Replacement Item:
This antibody may replace item sc-166787 from Santa Cruz Biotechnology.
Description of Target:
ASL is a member of the lyase 1 family. The protein forms a cytosolic homotetramer and primarily catalyzes the reversible hydrolytic cleavage of argininosuccinate into arginine and fumarate, an essential step in the liver in detoxifying ammonia via the urea cycle. Mutations in its gene result in the autosomal recessive disorder argininosuccinic aciduria, or argininosuccinic acid lyase deficiency.This gene encodes a member of the lyase 1 family. The encoded protein forms a cytosolic homotetramer and primarily catalyzes the reversible hydrolytic cleavage of argininosuccinate into arginine and fumarate, an essential step in the liver in detoxifying ammonia via the urea cycle. Mutations in this gene result in the autosomal recessive disorder argininosuccinic aciduria, or argininosuccinic acid lyase deficiency. A nontranscribed pseudogene is also located on the long arm of chromosome 22. Alternatively spliced transcript variants encoding different isoforms have been described.
Protein Size (# AA):
444
Molecular Weight:
49kDa
Host:
Rabbit
Clonality:
Polyclonal
Purification:
Protein A purified
Application:
WB
Tissue Tool:
Find tissues and cell lines supported by DNA array analysis to express ASL.
RNA Seq:
Find tissues and cell lines supported by RNA-seq analysis to express ASL.
Immunogen:
The immunogen is a synthetic peptide directed towards the middle region of human ASL
Tested Species Reactivity:
Human
Predicted Homology Based on Immunogen Sequence:
Cow: 100%; Dog: 100%; Goat: 100%; Guinea Pig: 100%; Horse: 100%; Human: 100%; Mouse: 100%; Rabbit: 100%; Rat: 100%; Yeast: 93%; Zebrafish: 93%
Complete computational species homology data:
Anti-ASL (ARP41667_T100)
Peptide Sequence:
Synthetic peptide located within the following region: LILYCTKEFSFVQLSDAYSTGSSLMPQKKNPDSLELIRSKAGRVFGREDK
Product Format:
Liquid. Purified antibody supplied in 1x PBS buffer with 0.09% (w/v) sodium azide and 2% sucrose.
Reconstitution and Storage:
For short term use, store at 2-8C up to 1 week. For long term storage, store at -20C in small aliquots to prevent freeze-thaw cycles.
Concentration:
Batch dependent within range: 100 ul at 0.5 - 1 mg/ml
Protein Interactions:
WDYHV1; ASL; GINS4; NCDN; TUFM; PDHA1; NEDD8; HK1; GDI1; BAG3; OVGP1; HMOX1; FBP1; CSNK2A2; UBC; QARS;
Blocking Peptide:
For anti-ASL (ARP41667_T100) antibody is Catalog # AAP41667 (Previous Catalog # AAPP24311)
Datasheets/Manuals:
Printable datasheet for anti-ASL (ARP41667_T100) antibody
Target Reference:
Tanaka,T., (2002) Tohoku J. Exp. Med. 198 (2), 119-124
Publications:

Hu, Z. et al. Quantitative liver-specific protein fingerprint in blood: a signature for hepatotoxicity. Theranostics 4, 215-28 (2014). WB, Cow, Dog, Goat, Guinea Pig, Horse, Human, Mouse, Rabbit, Rat, Yeast, Zebrafish 24465277

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02/01/2017 16:23
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Product Review: ASL antibody-middle region (ARP41667_T100) in COS-7 using Western blot
Product Page for ASL antibody-middle region (ARP41667_T100)

Researcher: Shawn Elms. Georgia Health Science University
Application: Western blotting
Species + Tissue/Cell type: Lane1: 10 ug COS-7 cell lysate
Primary antibody dilution: 1:1000
Secondary antibody: Anti-rabbit HRP
Secondary antibody dilution: 1:2000


Questionnaire:
How do Aviva's reagents play a role in your experimental goals? We attempted to probe for endogenous ASL in endothelial cells and endogenous as well as overexpressed COS-7 cells
How would you rate this antibody on a scale from 1-5 (5=best) and why? 4. I could get a clean blot at 49.
Would you use this antibody in future experiments? Yes
Have you used another antibody which has worked in your application? No
Do you believe the information about the reagent on Aviva's website is correct? Yes
If the antibody works, do you plan to use it in future experiments or to publish your data? Why or why not? Yes. I was able to get a clean ASL blot
How did you store the antibody after re-suspension? At -20
Sample Description (please include 1) species type, and 2) cell/tissue type, and 3) how much protein loaded for each lane of your gel): COS-7 cell culture (fibroblast); 10ug per lane
How many different experimental trials were conducted using the antibody sample? 6
How was this sample prepared? COS-7 cells were transfected with ASL pcDNA or native BAECs were grown to confluence. After 24 hours the cells are lysed in lysis buffer, sonicated, boiled for five minutes, spun down and run on a 10% gel for Western blotting.
Primary antibody dilution and incubation time: 1ug/ml. Overnight
Secondary antibody used and dilution and incubation time: Rabbit 0.5ug/ml. one hour
What controls were used in your experiment (positive/negative)? Non-transfected COS-7 and bovine aortic endothelial cells were both used as positive cells
Please include your detailed WB Procedure/Protocol here: Make 10% APS freshly in an eppendorf tube and vortex it, which is 0.1g APS in 1 ml deionized water.Use 50 ml tubes to make lower and upper gels.Lower gel
10% gel  7%gel   Upper gel
Acrylamide     5ml  3.5ml      1.3ml
Lower buffer 3.75ml  3.75ml  Upper b 2.5ml
Water            6.25ml  7.75ml      6.05ml
10%APS 75ul  75ul       75ul
TEMED 15ul  15ul             15ul

Pour 4.5 ml lower gel to each space (for 1 mm) and 7.5ml (for 1.5 mm), fill the top with deionized water. Wait for 10 min until gel is solidified.

Aspirate away the water, use filter paper to wipe the upper part of the well.

Get the upper gel ready, fill the upper part of the well. Then put the comb in it.

Wait for solidification of upper gel (around 5 min)

Make running buffer 500 ml for 2 gel (100 ml 5* running buffer and 400 ml water)

Take the gels out and remove the combs, put them on the cast, big glass towards the outside, fasten the clips.

Put the cast into the box and pour running buffer into it, first fill the space between the two gels in the middle.

Spin the thawed samples for 8 min (14000R 4 degree)

Take 100 ul supernatant of each sample, add 20ul 6* sample buffer to each well, vortex them.

Use needle to punch a hole on the lid of each tube, boil samples for 5 min, quick spin again for 1 min.

Loading the samples 20ul each into the gel, put the syringe into the bottom of the well and load slowly. The first well should be MW standard for 5ul in the freezer.

Set 70mV for running, after the first blue ladder comes out, (about 15-20 min) change the voltage to 120mV, wait until the blue line comes out of the bottom line. In this way we can prevent deformation of the gel. (takes 2 hrs total)

For transfer buffer, we can use 10* transfer buffer 100ml, add water to 800ml, mix them then add 200ml methanol.
(For transfer buffer - 3.02 Tris, 14g glycine, 800ml water. After they are mixed, add 200ml methanol)

For each gel, use 2 sponge and 4 filter paper, cut the membrane at the same size with the filter paper. Soak all these stuff I the transfer buffer.

Make a sandwich, first drop some buffer onto the gel, be sure not to leave the gel dry. Use a hollow tube to drive the bubbles away.

Black to back of the frame, put a box of ice into it, when it's connected with the lid, remember black to black, red to red.

Transfer at 45mV overnight in the fridge. If it's in a hurry, we can the transfer at 100mV for 1 hr.

To test if transfer is good enough:
1. Use coomassie blue to stain the gel, then use coomassie destain buffer to wash. After that we may check the gel if there is a lot of protein on it.
2. Use Ponceao Red to stain the membrane then wash with water, we can find a lot of protein stuck on the membrane.

Next day, take out the membrane, wash with TBS-T, block with 5%milk (2.5g milk in 50ml TBS-T) for 30-60 min.

Incubate with 1Ab for 30 -60 min, wash with TBS-T for 5min * 2

Incubate with 2Ab for 30 -60 min, wash with TBS-T for 5min * 4, then 5min * 2

Blend reagent A&B 1:1 and incubate the membrane for 1 min, fix the membrane in the cassette and develop the film, label the blots.
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