Rabbit anti-Goat IgG (Whole Molecule) Antibody (Colloidal Gold (5nm))

Details for Product No. ABIN3042038, Supplier: Log in to see
Antigen
Epitope
Whole Molecule
4684
2572
1882
1039
1015
825
305
273
35
34
31
8
7
7
3
3
2
2
1
1
1
1
1
1
Reactivity
Goat
2780
2760
2347
1724
1435
778
607
426
364
338
330
324
307
290
169
148
126
114
41
29
27
25
12
11
9
8
6
5
5
4
4
4
4
4
4
4
4
3
3
3
2
2
2
1
1
1
1
1
Host
Rabbit
6750
4660
1834
812
684
394
88
52
51
9
8
5
5
4
2
1
1
Clonality
Polyclonal
Conjugate
Colloidal Gold (5nm)
2183
1862
1718
1362
765
675
521
297
295
284
274
268
217
186
178
117
102
79
69
58
54
40
35
26
23
22
19
19
19
19
19
16
15
15
15
15
15
15
14
13
12
11
11
10
10
9
9
9
9
9
9
9
8
8
8
7
6
6
6
6
6
6
6
5
5
5
5
5
4
4
4
4
3
3
3
3
3
3
3
3
3
2
2
2
2
2
2
2
2
2
2
2
2
2
1
1
1
1
1
1
1
1
Application
Immunoelectron Microscopy (IEM), Immunocytochemistry (ICC), Immunohistochemistry (Frozen Sections) (IHC (fro)), Immunohistochemistry (Paraffin-embedded Sections) (IHC (p))
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'Independent Validation' Badge
Antigen Rabbit IgG
Method validated Electron Microscopy
Positive Control Lab stock CBD-SNAP antibody
Negative Control No SNAP-tag antibody
Notes We validate that the anti-Rabbit IgG (Whole Molecule) Antibody (Colloidal Gold (5nm)) ABIN3042038 specifically a rabbit primary antibody in SEM-EBSD images of oyster visceral mass tissue.
Primary Antibody ABIN1573927
Secondary Antibody ABIN3042038
Protocol
  • Oyster visceral mass tissue is dissected and fixed in 4% paraformaldehyde in seawater overnight.
  • Sections are rinsed with 70% ethanol and then transferred to 70% ethanol for about 2h at RT before automated dehydration process.
  • Serial dehydration process using an automated ASP300S Enclosed Tissue Processor (Leica Biosystems) as follows:
    • 70% ethanol for 45min
    • 90% ethanol twice for 45min
    • 100% ethanol twice for 45min
    • xylene twice for 45min
    • paraffin wax at 58°C 3 times for 30min
  • Tissue is mounted in a paraffin block and hardened overnight.
  • 8µm tissue sections are retrieved from the block and collected on circular glass cover slips.
  • Heat cover slips at 60°C for 1h.
  • Deparaffination and rehydration:
    • xylene twice for 15min
    • 100% ethanol twice for 10min
    • 95% ethanol for 10min
    • 85% ethanol for 10min
    • 70% ethanol for 10min
    • 50% ethanol for 10min
    • 30% ethanol for 10min
    • distilled water for 10min
    • PBS for 10min
  • Wash tissue sections with PBS with 0.05% triton X twice for 30min.
  • Permeabilize in PBS with 0.05% triton X overnight.
  • Treatment of the tissue sections with 1mg/mL sodium borohydride in PBS three times for 5min to reduce autofluorescence.
  • Wash sections in PBS 3 times for 15min for at RT.
  • Block sections in PBST with 1% BSA for 2h at RT.
  • Incubate sections with CBD-SNAP antibody (lab stock) diluted 1:200 in PBST with 1% BSA overnight at 4°C to detect the location of chitin.
  • Wash sections in PBS 3 times for 15min with PBS at RT.
  • Additionally, incubate the CBD-SNAP and SNAP-tag double-stained sections with rabbit anti-SNAP antibody (antibodies-online, ABIN1573927, lot 13D000621) diluted 1:200 in PBST with 1% BSA overnight at 4°C.
  • Wash sections in PBS 3 times for 15min at RT.
  • Incubate sections with the secondary rabbit anti-rabbit IgG (whole molecule) antibody Colloidal Gold (5nm) conjugate (antibodies-online, ABIN3042038) diluted 1:50 in PBST with 1% BSA in the dark overnicht at 4°C.
  • Wash sections in PBS three times for 15min at RT.
  • Post fixsections with 1% glutaraldehyde in PBS for 15min at RT.
  • Wash sections in distilled water three times for 10min at RT.
  • R-Gent SE-EM silver enhancement reagents (Aurion, 2551, lot 60323/2):
    • Prepare a slower developer mix with 1:60 ratio of initiator:activator.
    • Silver enhancement for 30 min with 1:20 developer: enhancer ratio.
    • Stop the reaction by washing with distilled water for 5min 3 times.
  • Graded alcohol series: /li>
    • 50% alcohol for 15min
    • 70% alcohol for 15min
    • 85% alcohol for 15min
    • 95% alcohol for 15min
    • 100% alcohol for 15min
  • Critical point drying of the sections with CO2 transitional fluid for 15min.
  • Slowly degas sections at a degassing rate of 100 Pa/min.
  • Sections are mounted on the same aluminum stub, coated with carbon and visualized at 20kV with probe current at 40, with basic SEM-EBSD mode on a Hitachi S-3400 Variable Pressure SEM, working distance kept at 10mm. Brightness and contrast settings are kept constant for the both samples.
Experimental Notes To validate the specificity of the anti-Rabbit IgG (Whole Molecule) Antibody (Colloidal Gold (5nm)) ABIN3042038, 8µm paraffin sections of oyster’s visceral mass were observed in this study. We compared the level of silver aggregating ability in the presence and absence of ABIN3042038, using commercially available silver enhancement method and SEM-EBSD imaging. We found that the test samples treated with anti-Rabbit IgG (Whole Molecule) Antibody (Colloidal Gold (5nm)) showed distinctive silver aggregates while the primary rabbit anti-SNAP antibody ABIN1573927 was present. These aggregates were not observable in the negative control.
Validation Images
Electron Microscopy validation image for Rabbit anti-Goat IgG (Whole Molecule) antibody (Colloidal Gold (5nm)) (ABIN3042038) SEM-EBSD images of oyster visceral mass tissue stained with CBD-SNAP and anti-SNAP pr...
Immunogen Goat IgG (whole molecule)
Isotype IgG
Specificity This antibody is specific for goat IgG
No Cross-Reactivity Human, Rat (Rattus), Mouse (Murine), Rabbit
Cross-Reactivity (Details) This colloidal gold conjugated antibody is specific for goat IgG and shows no cross-reactivity with human/ rat/mouse/rabbit IgG.
Purification This antibody is purified from antiserum by immunoaffinity chromatography which removes essentially all rabbit serum proteins, except the specific antibody for goat IgG. The rabbit anti-goat IgG is conjugated to 5 nm colloidal gold and generates electron dense particles that are visible using electron microscopy.
Research Area Immunology, Secondary Antibodies
Application Notes Electron Microscopy|1:20-50| Immunohistochemistry(Paraffin-embedded Section)|1:50-100| Immunohistochemistry(Frozen Section)|1:50-100| Immunocytochemistry|1:50-100|
Restrictions For Research Use only
Format Liquid
Handling Advice Do not freeze.
Storage 4 °C
Storage Comment At 4°C for one year.
Expiry Date 12 months