Blood Culturing

Defining bacteremia is a very important part of microbiologic investigation in hospital and many misconceptions exist about the practice. The following gives practical guidance.


Blood Cultures Step by Step

First Venipuncture

Prepare the skin overlying the selected vein thoroughly.

  • Apply chlorhexidine in alcohol antiseptic solution and allow to stand for at least 1 minute before proceding.
  • Don’t palpate the vein after you have disinfected the skin!!

Two methods of venipuncture are acceptable:

  • Needle and syringe +/- butterfly
  • Butterfly and adapter with vacutainer barrel – This is the preferred method as the needle can be secured leaving both hands free for subsequent manipulations. Also the Bactec bottles can be kept upright allowing for proper level of fill. After the cultures are completed other tubes can be drawn for hematology, chemistry etc.

Clean the bottle septa with alcohol swab.

  • Do not use povidone – iodine. Draw 20 mls of blood for culture and divide equally (i.e. 10 ml per bottle) between the aerobic (resin containing white topped) bottle and the anaerobic (lavender topped) bottle. Avoid under or overfilling. If less than 10 ml can be collected place it in the aerobic bottle only.

Label the bottles.

  • If using an identification sticker DO NOT COVER THE BAR CODE on the bottle
  • Identify the site (e.g. right antecubital fossa)
  • Minimum patient identification must include Name and Hospital number

If the bottle is improperly labeled it will not be processed in the lab.

Fill in the requisition.

Arrange for prompt transport to laboratory.

  • During lab hours bring the bottles directly to the lab
  • After hours place the bottles in the incubator

Second Venipuncture

Immediately after the first venipuncture, proceed with the second venipuncture at a separate site.

Follow the exact procedures outlined above except draw only ten ml for culture and place it all in an aerobic (white topped) bottle (no anaerobic bottle). Remember, this second venipuncture requires a second requisition.


Important Variables in Blood Culturing


This is, without question, the most important consideration. Generally, the higher the volume that is cultured the higher the yield of microorganisms. Of course, there is an upper limit to sampling of blood for culture and when all factors are weighed, 30 – 40 ml per blood culturing episode is the optimum. Greater than 95% of isolates will be found in the first 40 mls of blood.


There is no controversy about one thing – one blood culture is never appropriate! The reason for this is that it is impossible to judge the probability of contamination with organisms on the skin. Also it is difficult to obtain sufficient volume with one venipuncture especially if using needle and syringe. Therefore two blood cultures (i.e. two venipunctures) is the minimum and in most cases is the optimum. In the case of suspected endocarditis order three or four cultures spaced in time to provide evidence of continous bacteremia.

How many in one day?

Two blood cultures will document the bacteremia in the vast majority of “septic episodes”. The majority of patients will be well served by having only their initial two blood cultures of any day. A few patients warrant culturing twice in one day (i.e. two cultures at one time and two cultures later in the day). It is the extremely rare patient that derives benefit from more than this. Of course these are statistical guidelines and clinical judgment must ultimately prevail.

How many in an admission?

In the case of fever of unknown origin or febrile neutropenia, if blood cultures drawn on three different days of the admission have not revealed an organism it is very likely that further conventional cultures will also be fruitless. Consideration to special cultures (e.g. fungal) and other diagnostic tests should be given. I have seen a bone marrow transplant recipient who had 47 blood cultures of 20 ml done during one admission!


This is the most controversial variable and the experts are divided into two camps:

  1. Bacteremia peaks before fever and therefore one should wait a period of time after the first blood culture in an attempt to find the next bacteremic peak. Times ranging from 20 minutes to several hours are advocated.
  2. Patients who need blood cultures often need empirical antibiotic therapy and therefore all the blood should be drawn at one time allowing expeditious therapy. Besides one time is as likely as any other to be a bacteremic time.

I am squarely planted in the second camp because it is the most practical. The only clinical circumstance that requires spacing of blood cultures is subacute bacterial endocarditis because of the need to prove continuous bacteremia.

Culture from Indwelling Lines

In general, lines of all types should be avoided. In any case of suspected “line sepsis” the first thing one must prove is the sepsis part – i.e. that the patient is bacteremic. If sampling is done both through the line and peripherally and only the line sample is positive it may reflect colonization of the line. This finding is difficult to interpret and only makes clinical decisions harder. It may prompt removal of a line that was not the source of the problem. If the line is cultured it must always be accompanied by a peripheral culture.

Blood Culture Protocol