Donkey Anti-Guinea Pig IgG/PE-CY5

The Donkey Anti-Guinea Pig IgG/PE-CY5 is a versatile antibody conjugate suitable for various applications, including Immunofluorescence (IF) and Flow Cytometry (FCM). Below are the protocols for each validated application.

For IF, the following steps are recommended:

- Reagents or Buffers: Phosphate-buffered saline (PBS), 4% paraformaldehyde for fixation, 0.1% Triton X-100 for permeabilization, and a mounting medium with DAPI for nuclear staining.

- Sample Preparation: Fix cells or tissue sections with 4% paraformaldehyde, permeabilize with 0.1% Triton X-100, and block with a suitable blocking buffer.

- Antibody Incubation: Dilute the Donkey Anti-Guinea Pig IgG/PE-CY5 1:100 to 1:1000 in the blocking buffer and incubate for 1-2 hours at room temperature or overnight at 4°C.

- Washing or Detection: Wash samples thoroughly with PBS, and then mount with a DAPI-containing mounting medium for nuclear counterstaining.

- Technical Notes: Optimize the antibody dilution based on the specific experimental conditions and the intensity of the fluorescence signal.

For FCM, follow these guidelines:

- Reagents or Buffers: PBS, FACS buffer (PBS with 1% BSA and 0.1% sodium azide), and a fixation buffer if necessary.

- Sample Preparation: Prepare single-cell suspensions and block with FACS buffer.

- Antibody Incubation: Dilute the Donkey Anti-Guinea Pig IgG/PE-CY5 1:100 to 1:1000 in FACS buffer and incubate for 30 minutes to 1 hour on ice.

- Washing or Detection: Wash cells with FACS buffer, and then analyze by flow cytometry.

- Technical Notes: Ensure proper compensation for the PE-CY5 channel, and consider using a fixation step if analyzing intracellular targets.

Both IF and FCM protocols require optimization of the antibody dilution and incubation times based on the specific experimental setup and the intensity of the signal obtained.

Use assay conditions validated for IF, apply Donkey Anti-Guinea Pig IgG/PE-CY5 at the recommended working dilution, include proper positive and negative controls, perform thorough wash steps, and document incubation time, temperature, and detection conditions for reproducible performance. Recommended dilution: FCM=1:100-1000;

IF=1:100-1000.