RHOT1 antibody (N-Term)

Catalog No. ABIN405704

Product Details anti-RHOT1 Antibody

Target: RHOT1 (Ras Homolog Gene Family, Member T1 (RHOT1))

Binding Specificity: N-Term

Reactivity: Human, Mouse, Rat, Dog, Cow, Guinea Pig, Horse, Rabbit, Zebrafish (Danio rerio)

Host: Rabbit

Clonality: Polyclonal

Conjugate: This RHOT1 antibody is un-conjugated

Application: Western Blotting (WB)

Sequence: EMKPACIKAL TRIFKISDQD NDGTLNDAEL NFFQRICFNT PLAPQALEDV

Predicted Reactivity: Cow: 100%, Dog: 100%, Guinea Pig: 100%, Horse: 100%, Human: 100%, Mouse: 100%, Rabbit: 100%, Rat: 100%, Zebrafish: 85%

Characteristics: This is a rabbit polyclonal antibody against RHOT1. It was validated on Western Blot using a cell lysate as a positive control.

Purification: Affinity Purified

Immunogen: The immunogen is a synthetic peptide directed towards the N terminal region of human RHOT1

Application Details

Application Notes: Optimal working dilutions should be determined experimentally by the investigator.

Comment:

Antigen size: 659 AA

Restrictions: For Research Use only

Independent Validation (WB)

Validation #100008 (Western Blotting)

by: Shaw Lab, Department of Biochemistry, University of Utah

No.: #100008

Date: 09/12/2015

Antigen: RHOT1

Method validated: Western Blotting

Positive Control:

Purified recombinant Miro1

Isolated mitochondria from immortalized Miro1 KO MEFs expressing Myc-Miro1

Negative Control:

Purified recombinant Miro2

Isolated mitochondria from immortalized Miro1 WT MEFs

Isolated mitochondria from immortalized Miro1 KO MEFs expressing Myc-Miro2

Notes:

Passed. ABIN405704 specifically recognizes Miro1 without cross-reaction with Miro2.

Validation Images

Full Methods

Primary Antibody: ABIN405704

Secondary Antibody: Goat-anti-rabbit IgG HRP conjugated (Sigma, A0545-1M22)

Full Protocol:

• Transfect immortalized Miro1-/- MEFs with Myc-Miro1-V1 or Myc-Miro2-V1 as described in PMID 25136135.
• Isolate mitochondria from immortalized Miro1+/+, Miro1-/-, Miro1-/- + Myc-Miro1 and Miro1-/- + Myc-Miro2 MEFs.
• Determine protein concentration and snap freeze mitochondria in liquid nitrogen.
• Thaw 30 µg mitochondria per lane on ice.
• Thaw 100 ng recombinant Miro1 and Miro2 per lane on ice.
• Add 2x Laemmli buffer and 2% BME to samples and denature for 5 min at 95 °C.
• Separte 30 µg/lane mitochondria and 100 ng/lane recombinant protein on a 12.5% SDS-PAGE for 3 h at 40mA/180V.
• Semi-dry transfer to PVDF membrane for 2 h at 250 mA/15 V.
• Block the membrane with TBST + 5% non-fat dry milk for 1 h at RT.
• Incubate with primary
• • rabbit anti-RHOT1 antibody (antibodies-online, ABIN405704) diluted 1:1000 in TBST + 3% BSA overnight at 4°C.
  • rabbit anti-Myc1 antibody diluted 1:1000 in TBST + 3% BSA overnight at 4°C.
  • rabbit anti-Cyt c antibody diluted 1:1000 in TBST + 3% BSA overnight at 4°C.
• Wash membrane 3x for 5 min with TBST buffer.
• Incubate with secondar goat anti-rabbit IgG HRP antibody (Sigma, A0545-1M2) diluted 1:2000 in TBST + 5% non-fat dry milk for 1 h at RT.
• Wash membrane 3x for 10 min with TBST buffer.
• Reveal protein bands on a Bio-Rad imaging system using homemade ECL prepared according to PMID 17141265.
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Handling

Format: Liquid

Concentration: Lot specific

Buffer: Liquid. Purified antibody supplied in 1x PBS buffer with 0.09 % (w/v) sodium azide and 2 % sucrose.

Preservative: Sodium azide

Precaution of Use: This product contains Sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.

Handling Advice: Avoid repeated freeze-thaw cycles.

Storage: -20 °C

Storage Comment: For short term use, store at 2-8°C up to 1 week. For long term storage, store at -20°C in small aliquots to prevent freeze-thaw cycles.

Target Details for RHOT1

Target: RHOT1 (Ras Homolog Gene Family, Member T1 (RHOT1))

Alternative Name: RHOT1 (RHOT1 Products)

Synonyms: rhot1 antibody, zgc:55581 antibody, zgc:77063 antibody, wu:fd14e11 antibody, wu:fi14a05 antibody, arht2 antibody, miro-2 antibody, rasl antibody, ARHT1 antibody, MIRO-1 antibody, MIRO1 antibody, 2210403N23Rik antibody, AA415293 antibody, AF244542 antibody, AI834919 antibody, Arht1 antibody, C430039G08Rik antibody, Miro1 antibody, miro1 antibody, si:dkeyp-97g3.6 antibody, ras homolog family member T1a antibody, ras homolog family member T2 antibody, ras homolog family member T1 antibody, ras homolog family member T1 L homeolog antibody, rhot1a antibody, rhot2 antibody, RHOT1 antibody, rhot1 antibody, rhot1.L antibody, Rhot1 antibody, rhot1b antibody

Background: RHOT1 is mitochondrial GTPase involved in mitochondrial trafficking. It is probably involved in control of anterograde transport of mitochondria and their subcellular distribution.
Alias Symbols: ARHT1, FLJ11040, FLJ12633, MIRO-1, MIRO1
Protein Interaction Partner: PCSK9, ERLIN2, PARK2, ILK, UBC, CFTR, PINK1, UBD, IRAK1, AMFR,
Protein Size: 659

Molecular Weight: 75 kDa

Gene ID: 55288

NCBI Accession: NM_001033566, NP_001028738

UniProt: Q8IXI2