No Staining

Secondary antibody is not compatible with primary antibody (always use antimouse for mouse, anti-rabbit for rabbit, etc.). 

Chromogenic substrate has been replaced with another that is not intended for use with the enzyme substrate. 

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No staining at all or very slight partial membrane staining in less than 10% of tumor cells. 

Storing and titrating antibodies correctly are often easy solutions when no staining is observed. Some antibodies are extremely sensitive to repeat freeze-thaw cycles and others have a limited shelf life, as evidenced by a short expiry dates, ft is important to check the compatibility of the primary and secondary antibodies before starting IHC, as using the wrong secondary IgG is a common but easily corrected problem. As mentioned, antigen retrieval can be a problem when tissue has been overfixed, so special attention should be paid to fixation time and, once optimized, should be held constant for all subsequent runs. 

Safety No stain should contain a hazardous material such as radioisotopes, cyanide,... 

Good charcoal is very dark, possesses a bright lustre and somewhat conchoidal fracture it resists gradual pressure to a considerable extent, and produces a sharp sonorous sound when allowed to fell upon a hard body. It should burn when ignited without either flame or smoke, and when handled no stain ought to remain. Although in bulk it floats in water,... 

With some experience, no staining of the membrane is necessary. When the PVDF membrane is illuminated with white light immediately after transfer, protein bands appear as white, opaque areas surrounded by translucent protein-free membrane. Taking care not to dry the membrane, a protein band to be eluted can be marked with a pencil. 

If no staining is observed in the positive control, check whether a reagent has been inadvertently omitted or whether the wrong reagent has been used (for example, antimouse Ig on a rat monoclonal). This is easy to do if several different antibodies from different species are being used. 

Fig. 6.2 HNE immunoreactivity in the hippocampus. A section from a normal rat showing no staining in field CA1 and the dentate gyrus (asterisks). B section from a 1 day post-kainate injected rat, showing dense staining in pyramidal cell bodies (arrows), and the neuropil (asterisk), in an affected area. Abbreviations CA, field CA1. DG, dentate gyrus. Scale = 250 /xm. Reproduced with kind permission from Ong et al., 2000b, Free Radical Biology and Medicine 28 1214-1221. Elsevier...

Fig. 39A, B Staining for BrdU in septal and caudate SVZa. A Control monkeys on days 9 and 23 after sham operation. B Postischemic monkeys on various time-points after ischemia. Note the increase of positive cells after ischemia. Clusters depicted by arrows are magnified in insets. The position of the visual field within the frontal lobe is schematically depicted as a. frame on the map (upper right). (-), negative control showing no staining for BrdU F, frontal cortex WM, white matter S, striatum. Asterisk, anterior horn of lateral ventricle. Scale bar= 100 pm...

Figure 19.1 Or85e is selectively expressed in subsets of maxillary palp neurons. In situ hybridization with an antisense Or85e probe reveals no staining in antenna (left) or brain (center). Nine cells stain in this section of the maxillary palp (right). The relative positions of the two olfactory sensory organs on the head of the adult fly are indicated with white dashed lines.

Untreated PEUU controls (which had been exposed to GOx) exhibited no stain after continuous stirring in SDS, Triton, and sodium phosphate. GOx-PPNVP/PEUU samples washed by the same method as well as PEUU controls washed by less stringent methods exhibited positive stain. Thus the immunochemical stain assay demonstrated that the continuous wash in SDS, Triton, and sodium phosphate removed physically adsorbed GOx from the surface of GOx-PPNVP/PEUU, leaving covalently bound GOx. Positive stain was easily observable with the naked eye, making the immunochemical stain an effective novel technique for the quick screening of wash procedures for thin film samples. 

An ideal negative cell line control will contain an amount of target antigen, sufficiently low to produce no staining if the procedure has been performed correctly. At the same time, the amount should be sufficiently high to produce a weakly positive stain if the run has been performed under conditions that produce an excessively strong stain. 

Little or no staining of controls or specimen tissue, except for counterstain. May show little or no background staining. 

NO STAINING SEEN. GO TO NEXT STEP. Block with levamisole (Intestinal alkaline phosphatase may be... 

Figure 10.4 FcyRl in human placenta. CD64 staining of Hofbauer cells and perivascular monocyte/macrophages and dendritic cells. No staining of placental trophoblast epithelium or endothelium. See color insert.

Figure 10.5 (a) Unexpected broad-spectrum cross-reactivity with epithelium, endothelium and selected vascular smooth myocytes. No staining of interstitial (stromal) cells, collagen, or nuclei. (b) No staining in replicate sections stained by negative control antibody at similar staining concentration. See color insert. 

Figure 10.9 Unexpected cross-reactivity with contractile filaments (cross-striations, intercalated discs) in cardiac myocytes. No staining of sarcolemmal cells, interstitial cells or endothelium. See color insert.

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