| No. | Specificity | Hybridoma | Isotype | Comments; % Saturation | Notes |
| 1 | Control | M1/69.HK | Rat IgG2b | Shows unwanted binding; n.d. | |
| 2 | CD4 | GK1.5 | Rat IgG2b | Use #A as 2nd Ab; 67% | |
| 3 | CD4 | H129.19 | Rat IgG2a | PE-conjugate. See * below.; 50% | |
| 4 | CD8 | M5/24 | Rat IgG2a | Use #A as 2nd Ab; ~50% | |
| 5 | CD8 | 53.6 | Rat IgG2a | PE-conjugate. See * below. 17% | |
| 6 | CD11a (LFA-1)§ | M17/7 | Rat IgG2b | Use #A as 2nd Ab. See § below; ~75% | |
| 7 | CD18 (LFA-1, CR3)§ | M18/2 | Rat IgG2a | Use #A as 2nd Ab. See § below; ~75% | |
| 8 | CD45/1 | M1/9.3 | Rat IgG2a | Use #A as 2nd Ab; 20% | |
| 9 | CD45/2 § | M1/89.18 | Rat IgG2b | Use #A as 2nd Ab. See § below; 33% | |
| 10 | IgG, mouse | Polyclonal | Goat mixed | FITC-conjugated; | |
| 11 | IgM, mouse | Polyclonal | Goat mixed | PE-conjugated; 33% | |
| 12 | ICAM-1, CD54 | YN1/1 | Rat IgG2a | Use #A as 2nd Ab; 67% | |
| 13 | MHC Class I, H-2KDL | M1/42 | Rat IgG2a | Use #A as 2nd Ab; 33% | |
| 14 | MHC Class II, H-2I-A, I-E | M5/114 | Rat IgG2b | Use #A as 2nd Ab; n.d. | |
| 15 | TCR | H57-597 | Hamster IgG | FITC-conjugated; 67% | |
| 16 | Thy-1 (mouse T cell marker) | M5/49 | Rat IgG2a | Use #A as 2nd Ab; 4% | |
| Numbered Abs above are specific for a marker on mouse lymphocytes. Lettered reagents below are for use as "2nd reagents" for indirect staining of numbered, marker-specific Abs which are not directly conjugated with a fluorochrome. | |||||
| A | IgG, rat | Polyclonal Goat | mixed | FITC-conjugated, see ‡ below; ~50% | |
Colors: FITC emits green light, PE emits red light. Unless indicated in the Comments column, the above antibodies are not directly conjugated with a fluorochrome. Only green (FITC) indirect stain (anti-rat IgG) is available for unconjugated rat monoclonal Abs. If you stain two markers in the same tube, they must use different colors.
% Occupancy: The percentage of receptors on the cell surface which will be occupied by antibody at the dilution provided for your use. n.d. = not determined. Monoclonal antibody binding is well-described by the one-site binding equation, I/Imax = C/(C + Ch), where I is the observed fluorescence intensity, Imax is the intensity at saturation, Ch is the concentration of antibody which half-saturates, and C is the concentration applied. This equation has been fitted to I vs. C data for most of these antibodies, thereby determining Imax and Ch, and allowing the % occupancy to be calculated at a given C. In two-color samples, green fluorescence intensities exceeding 100 times autofluorescence make it difficult to get accurate red fluorescence readings (the compensation subtraction of green spill-up into the red channel fails). Therefore, green-reported antibodies specific for very high-density receptors (e.g. Thy-1, CD45) are intentionally diluted to give a low % saturation.
* If you use a rat PE conjugate in the same tube with an unconjugated rat Ab to a different marker, the PE conjugate must be applied after the 1st rat Ab and 2nd FITC anti-rat Ab, as a 3rd cycle. This avoids unwanted green staining of the red rat PE conjugate by the green anti-rat 2nd Ab.
‡ Use as 2nd Ab (indirect stain) for all unconjugated rat monoclonal 1st Abs. Absorbed with mouse Ig, so will not stain mouse B cell Ig by cross reaction.
§ Although 100% of splenocytes are positive with these antibodies, they stain T and B cells with different intensities. (The other antibodies above which stain 100% of splenocytes stain them all with a uniform intensity, namely the other CD45 and MHC class I.)