7.4.1 Gently shake incubated sample mixture and transfer 1 mL to 9 mL tempered (35-42°C) tetrathionate brilliant green broth (TBG). Mix inoculated broth on vortex mixer or by hand to disperse inoculum. Incubate for 7 h at 42°C (preferably in a water bath).

7.5.1 Each test unit has 2 chambers: inoculation chamber and motility chamber (Figure 1). Each step of the preparation sequence can be performed on an individual unit or multiple units as needed. Sample nos. can be recorded on lower portion of motility chamber, but must not interfere with reading of results. Alternatively, sample nos. may be recorded on flat surface of white cap.

7.5.2 Position unit with white cap UP, and remove white cap. Snip off tip of gel-former plug with alcohol-sterilized shears or clippers and discard tip (Figure 2). Cut should be made at point where tip meets base of plug. If tip of gel-former plug is not removed, antibody solution will be displaced from gel void when white cap is replaced. If water bubbles or pieces of gel block the opening, carefully insert a sterile Pasteur pipet into the gel void and aspirate any material.

7.5.3 Add 1 drop of antibody preparation to gel void in motility chamber (Figure 3). Replace white cap and screw on tightly. Antibody preparation should fill ca 2/3 of gel void. This can be detected by observing blue antibody solution in gel void.

7.5.4 For all samples, position unit with black cap UP, and remove black cap.

7.5.5 Remove chamber plug from inoculation chamber with sterile forceps and discard plug (Figure 4). If chamber plug is not removed, bacteria will be unable to move from inoculation chamber to motility chamber. If preparing 1-2 Test ahead of inoculation, replace black cap, Otherwise inoculate, as below.

Agitate incubated TBG to resuspend material. Aseptically pipette 1.5 mL of the TBG into the sidearm (empty inoculation chamber) of the 1-2 Test. Replace black cap tightly (Figure 5).

Place inoculated unit in incubator with white cap UP. Incubate unit in shipper/incubation tray at 35°C for 14-17 h.

7.8.1 With white cap UP, hold unit next to strong light. Desktop fluorescent light is recommended for reading test results. Carefully observe motility chamber gel by rotating unit back and forth through 360^o turn in front of light source.

7.8.2 Positive test is indicated by the presence of a white immunoband that is U-shaped or meniscus-shaped. Bands may be fully formed or may be more distinct on one side of the gel. Band is seen in upper half of motility chamber gel.

7.8.3 Positive test indicates that sample presumptively contains Salmonella. Positive test results should be confirmed.