Ras Homolog Gene Family, Member T1 (RHOT1) (N-Term) antibody

Details for Product No. ABIN405704, Supplier: Login to see New
New
Independently validated
Antigen
Synonyms rhot1, zgc:55581, zgc:77063, wu:fd14e11, wu:fi14a05, arht2, miro-2, rasl, ARHT1, MIRO-1, MIRO1, 2210403N23Rik, AA415293, AF244542, AI834919, Arht1, C430039G08Rik, Miro1, miro1, si:dkeyp-97g3.6
Epitope
N-Term
(24), (15), (3), (3), (2), (2), (2), (2), (2), (1)
Reactivity
Chicken, Cow (Bovine), Human, Mouse (Murine), Rat (Rattus), Zebrafish (Danio rerio)
(47), (14), (11), (4), (2), (2), (2), (2), (1), (1)
Host
Rabbit
(39), (8)
Clonality
Polyclonal
Conjugate
Un-conjugated
(2), (2), (2), (2), (2), (2)
Application
Western Blotting (WB)
(43), (23), (9), (2), (1)
Pubmed 1 reference available
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Quantity 50 μg
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Immunogen Synthetic peptide directed towards the N terminal of human RHOT1
Blocking Peptide Blocking peptide for this product available: Ras Homolog Gene Family, Member T1 (RHOT1) (N-Term) peptide (ABIN982970)
Sequence EMKPACIKAL TRIFKISDQD NDGTLNDAEL NFFQRICFNT PLAPQALEDV
Predicted Reactivity Chicken : 100 %, Dog : 100 %, Guinea pig : 100 %, Horse : 100 %, Pig : 100 %, Rabbit : 100 %, Rat : 100 %
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
Alternative Name RHOT1 (RHOT1 Antibody Abstract)
Background RHOT1 is mitochondrial GTPase involved in mitochondrial trafficking. It is probably involved in control of anterograde transport of mitochondria and their subcellular distribution.
Molecular Weight 75 kDa
Gene ID 55288
NCBI Accession NP_001028738, NM_001033566
UniProt Q8IXI2
Application Notes Optimal working dilutions should be determined experimentally by the investigator.
Comment

Antigen size: 659 AA

Restrictions For Research Use only
Format Lyophilized
Reconstitution Add 50 µL of distilled water.
Concentration 1 mg/mL
Buffer PBS buffer with 2 % sucrose
Handling Advice Avoid repeated freeze-thaw cycles.
Storage -20 °C
Storage Comment For longer periods of storage, store at -20 °C
Supplier Images
Western Blotting (WB) image for anti-Ras Homolog Gene Family, Member T1 (RHOT1) (N-Term) antibody (ABIN405704) WB Suggested Anti-RHOT1 Antibody Titration: 0.2-1 ug/ml
ELISA Titer:
Background publications Fransson, Ruusala, Aspenström: "The atypical Rho GTPases Miro-1 and Miro-2 have essential roles in mitochondrial trafficking." in: Biochemical and biophysical research communications, Vol. 344, Issue 2, pp. 500-10, 2006 (PubMed).

'Independent Validation' Badge
Antigen Ras Homolog Gene Family, Member T1 (RHOT1) (N-Term)
Method validated Western Blotting
Positive Control 100ng Recombinant Miro1; 30ug Mitochondria isolated from Miro1 WT MEFs and Miro1 KO MEFs expressing Myc-Miro1; in house purification and isolation Isolated mitochondria from immortalized Miro1 WT MEFs rMiro1, mtMiro1 wt MEFs, mtMiro1 KO MEFs plus Venus-Miro1
Negative Control 100ng Recombinant Miro2, 30ug Mitochondria isolated from Miro1 KO MEFs and Miro1 KO MEFs expressing Myc-Miro2; in house purification and isolation
Notes ABIN405704 is specific for Miro1 without cross-reaction with Miro2.
Primary Antibody ABIN405704
Secondary Antibody Goat-anti-rabbit IgG HRP conjugated (Sigma A0545-1M2 - 057K4802) 1:2000 in TBS + 0.05% Tween-20 + 5% non-fat dry milk
Protocol
  • Transfection of immortalized Miro1-/- MEFs with Myc-Miro1-V1 or Myc-Miro2-V1 (Published in Nguyen et al., 2014 (PNAS, 10.1073/pnas.1402449111))
  • Isolation of mitochondria from immortalized Miro1+/+, Miro1-/-, Miro1-/- + Myc-Miro1 and Miro1-/- + Myc-Miro2 MEFs
  • Determine protein concentration and snap freeze mitochondria in liquid nitrogen

    Day 1

  • Thaw 30ug mitochondria per lane on ice
  • Thaw 100ng recombinant Miro1 and Miro2 per lane on ice (in house purification)
  • Add 2x Laemmli buffer + 2% BME to samples and denature for 5 min at 95C
  • Load 30ug/lane mitochondria and 100ng/lane recombinant protein onto 12.5 % SDS-PAGE
  • Fermentas PageRuler Plus Prestained (2ul) - Lane 7 / 14 / 21
  • Recombinant Miro1- Lane 1 / 8 / 15
  • Recombinant Miro2- Lane 2 / 9 / 16
  • Miro1+/+ mitochondria- Lane 3 / 10 / 17
  • Miro1-/- mitochondria- Lane 4 / 11 / 18 / 22
  • Miro1-/- + Myc-Miro1-V1 mitochondria- Lane 5 / 12 / 19 / 23
  • Miro1-/- + Myc-Miro2-V1 mitochondria- Lane 6 / 13 / 20 / 24
  • Run gel 40mA 180V for 3h
  • Semi-dry transfer to PVDF 250mA 15V for 2h
  • Block with TBS + 3% BSA for 1h at RT
1st AB over night at 4C rocking
LanesAntibodyHostDilutionIn
1-6Anti-Miro1 Other company---
8-13Anti-Miro1 ABIN634527vrabbit1:1000TBST + 3% BSA
15-20Anti-Miro1 ABIN635090rabbit1:1000TBST + 3% BSA
22-24Anti-Myc---
All lanesAnti-Cyt c---
Anti-PRX3----

Day 2

  • 3x washing with TBS + 0.05% Tween-20 for 5'
  • 2nd AB for 1h at RT
  • Goat-anti-rabbit IgG HRP (Sigma A0545-1M2 - 057K4802)
  • 1:2000 in TBS + 0.05% Tween-20 + 5% non-fat dry milk3x washing with TBS for 10'
  • Detection with homemade ECL prepared according to Haan and Behrmann, 2007 (J Immunol Methods, 10.1016/j.jim.2006.07.027) on a Bio-Rad imaging system
Validation Images
Western Blotting validation image for anti-Ras Homolog Gene Family, Member T1 (RHOT1) (N-Term) antibody (ABIN405704) Western blot customer validation ABIN405704. Refer to the protocol section for more i...
Catalog No. ABIN405704
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