Mar 28, 2026
anti-MOR and anti-NeuN Immunocytochemistry
- Beatriz Aoyama1,2,
- Aashka Popat1
- 1UCSF;
- 2State University of Campinas
External link: https://doi.org/10.1002/cne.70175
Protocol Citation: Beatriz Aoyama, Aashka Popat 2026. anti-MOR and anti-NeuN Immunocytochemistry. protocols.io https://dx.doi.org/10.17504/protocols.io.8epv5kejdv1b/v1
Manuscript citation:
Popat AK, Simon RC, Aoyama BB, Wokhlu A, Ehrlich AT, Harwell CC, Margolis EB (2026) Mu Opioid Receptor mRNA and Protein Localization Across the Rat and Mouse Habenula. The Journal of Comparative Neurology 534(6). doi: 10.1002/cne.70175
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: September 12, 2025
Last Modified: March 28, 2026
Protocol Integer ID: 227077
Keywords: mu opioid receptor, opioid receptor, neuronal nuclei, mor, labeling mu
Abstract
Immunocytochemistry protocol for labeling mu opioid receptor (MOR) and neuronal nuclei (NeuN). Specifically used in rat and mouse tissue.
Protocol materials
Anti-NeuN Antibody, clone A60Merck MilliporeSigma (Sigma-Aldrich)Catalog #MAB377
Cy™5 AffiniPure® Goat Anti-Rabbit IgG (H L)Jackson ImmunoResearchCatalog #111-175-144
Fluorescein (FITC) AffiniPure® Goat Anti-Rabbit IgG (H L)Jackson ImmunoResearchCatalog #111-095-003
Cy™3 AffiniPure® Goat Anti-Mouse IgG (H L)Jackson ImmunoResearchCatalog #115-165-003
VECTASHIELD Antifade Mounting MediumVector LaboratoriesCatalog #H-1000
Bovine Serum AlbuminMilliporeSigmaCatalog #A3294
Normal Goat SerumJackson ImmunoResearchCatalog #005-000-121
ImmunoStar Mu Opioid Receptor AntibodyImmunostarCatalog #24216
Day 1
Wash slices 4x5 mins in 1xPBS on rotator at Room temperature
Incubate slices in blocking solution 500 µL per well , rotating for 2 hours at Room temperature
Blocking Solution:
- 0.2% BSA (0.2g/100mL)Bovine Serum AlbuminMilliporeSigmaCatalog #A3294
- 5% NGS (50 microliters/mL)Normal Goat SerumJackson ImmunoResearchCatalog #005-000-121
- 0.3% Tween20/1x PBS
Incubate slices in primary antibody solution 500 µL per well . Agitate overnight at 4 °C
Primary antibody solution:
- Rb anti-MOR 1:10000 ImmunoStar Mu Opioid Receptor AntibodyImmunostarCatalog #24216
- 0.3% Tween20/1x PBS
Day 2
Wash slices 5x10 mins in0.3%Tween20/1x PBS on rotator at Room temperature
Incubate slices in secondary antibody solution 500 µL per well , agitating overnight at 4 °C . Well plate should be covered with foil.
Secondary antibody solution:
- Cy5 Goat anti-Rb 1:500 for rat tissueCy™5 AffiniPure® Goat Anti-Rabbit IgG (H L)Jackson ImmunoResearchCatalog #111-175-144 , FITC Goat anti-Rb 1:500 for mouse tissue Fluorescein (FITC) AffiniPure® Goat Anti-Rabbit IgG (H L)Jackson ImmunoResearchCatalog #111-095-003
- 0.3%Tween20/1x PBS
Day 3
Wash slices 5x10 mins in0.3%Tween20/1x PBS on rotator at Room temperature
Incubate slices in primary antibody solution 500 µL per well , rotating for 3 hours at Room temperature
Primary antibody solution:
- mouse anti-NeuN 1:1000Anti-NeuN Antibody, clone A60Merck MilliporeSigma (Sigma-Aldrich)Catalog #MAB377
- 0.3% Tween20/1x PBS
Wash slices 4x10 mins in0.3%Tween20/1x PBS on rotator at Room temperature
Incubate slices in secondary antibody solution 500 µL per well , agitating for 1 hour at Room temperature
Secondary antibody solution:
- Cy3 Goat anti-Ms 1:500 Cy™3 AffiniPure® Goat Anti-Mouse IgG (H L)Jackson ImmunoResearchCatalog #115-165-003
- 0.3%Tween20/1x PBS
Wash slices 4x10 mins in0.3%Tween20/1x PBS on rotator at Room temperature
Wash slices 1x10 min in 1x PBS on rotator at Room temperature
Mount tissue onto slide with VectashieldVECTASHIELD Antifade Mounting MediumVector LaboratoriesCatalog #H-1000 and seal with nail polish.