Table 1.
Comparison of silver staining protocols from this method with others.
| Step | Method 1 Morrissey [4] |
Method 2 Bassam et al. [5] |
Method 3 Sanguinetti et al. [6] |
Method 4 Creste et al. [7] |
Method 5Qu et al. [8] | Method 6 Modification of Benbouza et al. [9] |
Method 7 Ji et al. [10] |
Method 8 Zhang et al. [11] |
Method 9 Han et al. [12] | Method 10 Byun et al. [13] | Method 11 An et al. [14] | Method 12 Our procedure |
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Prefix | 50% methanol, 10% acetic acid (10 min) 5% methanol, 7% acetic acid (30 min) |
– | – | 1.5% nitric acid (3 min) | – | – | – | – | – | – | – | – |
| Fixation | 10% glutaraldehyde (30 min) | 10% acetic acid(20 min) | 10% acetic acid 0.5% acetic acid(3 min) |
10% ethanol, 1% acetic acid (10 min) | – | 10% absolute ethanol 0.5% acetic acid (5 min) |
– | – | – | – | – | – |
| Rinse | H2O (overnight) H2O (30 min) H2O (2 h) |
H2O (2 min) 3 times | – | H2O (1 min) | – | – | – | H2O (10 s) Two times | – | – | – | H2O (5 s) 2 times |
| Soaking | 5g/ml dithiothreitol (30 min) | – | – | – | – | – | – | – | – | – | – | – |
| Impregnation | 0.1% AgNO3 (30 min) | 0.1% AgNO3 1.5 ml 37% HCOH (30 min) |
2% AgNO3 (5 min) | 2% AgNO3 (20 min) | 25% ethanol, 1% nitric acid, 2% AgNO3 (5–10) |
1.5% AgNO3, 1.5 ml 37% HCOH (22–24 °C) (6–7 min) |
0.1% AgNO3 (10–15 min) |
0.1% AgNO3 (4 °C) (15 min) |
1% nitric acid 0.1% AgNO3 (5 min) |
10% ethanol, 0.5%acetic acid, 0.2% AgNO3 (3–20 min) |
5% ethanol, 1% nitric acid, 0.1% AgNO3 (5 min) |
10% ethanol 0.7% nitric acid 0.22% AgNO3, (Room temp.) (5–30 min) |
| Rinse | H2O One time | H2O, 20 s, optional | Distilled H2O, 1 s | H2O (30 s) Two times |
H2O (3 min) | Distilled H2O, (10 s) 3 times | H2O (5 s) Two times |
2% NaOH, 0.01% HCOH, (10 s) |
H2O (5 s) Two times |
H2O (5 s) One times |
H2O (10 s) | – |
| Development | 50 l 37% formaldehyde + 100 ml 3% Na2CO3 3–5 min |
3% Na2CO3, 1.5 ml 37% HCOH; 2 mg Na2SO3.5H2O (10 °C) (2–5 min) |
1.5% NaOH, 2 ml 37% HCOH (5 min) |
3% Na2CO3, 1.5 ml 0.54 ml 37% HCOH (4-7 min) |
3% Na2CO3, 0.2% HCOH, (2–5 min) |
1.5% NaOH, 2 ml 37% HCOH (22–24 °C) (3–5 min) |
0.04% Na2CO3, 0.2% HCOH, (5–6 min) |
2% NaOH, 0.01% HCOH, (2-3 min) |
2% NaOH, 0.04% NaCO3, 0.0025% EBT, 0.15% HCOH (5 min) |
3% NaOH, 0.1% HCOH (55 °C) (5–10 min) |
1.3% NaOH, 0.65% NaCO3, 0.4% HCOH; (2–3 min) |
Per liter: 22.9 g Na2CO3, 1.25 ml 37% HCOH, 2 mg Na2SO3.5 H2O (4 °C) (3–5 min) |
| Stop | 5 ml of 2.3 M citric acid | 10% acetic acid(10 °C) (5 min) |
– | 5% acetic acid (5 min) |
10% acetic acid,(2–5 min) | 0.5% acetic acid 10% absolute ethanol (2 min) |
H2O (5 s) Two times | H2O (5 s) Two times |
2.5% ampicillin (5 s)Two times | 10% ethanol, 0.5%acetic acid (1 min) |
5% ethanol, 1% nitric acid (1 min) |
250 ml of 3% acetic acid (4 °C) 10% ethanol 5 min |